Spiro-lactam nmda receptor modulators and uses thereof

ABSTRACT

Disclosed are compounds having enhanced potency in the modulation of NMDA receptor activity. Such compounds are contemplated for use in the treatment of conditions such as depression and related disorders. Orally available formulations and other pharmaceutically acceptable delivery forms of the compounds, including intravenous formulations, are also disclosed.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. Ser. No. 14/764,402, filedJul. 29, 2015, which is a United States National Stage of InternationalApplication No. PCT/US2014/013621, filed on Jan. 29, 2014, which claimsthe benefit of U.S. Provisional Application No. 61/757,920, filed onJan. 29, 2013, each of which is incorporated by reference in theirentirety.

BACKGROUND

An N-methyl-d-aspartate (NMDA) receptor is a postsynaptic, ionotropicreceptor that is responsive to, inter alia, the excitatory amino acidsglutamate and glycine and the synthetic compound NMDA. The NMDA receptorcontrols the flow of both divalent and monovalent ions into thepostsynaptic neural cell through a receptor associated channel (Fosteret al., Nature 1987, 329:395-396; Mayer et al. Trends in Pharmacol. Sci.1990, 11:254-260). The NMDA receptor has been implicated duringdevelopment in specifying neuronal architecture and synapticconnectivity, and may be involved in experience-dependent synapticmodifications. In addition, NMDA receptors are also thought to beinvolved in long term potentiation and central nervous system disorders.

The NMDA receptor plays a major role in the synaptic plasticity thatunderlies many higher cognitive functions, such as memory acquisition,retention and learning, as well as in certain cognitive pathways and inthe perception of pain (Collingridge et al., The NMDA Receptor, OxfordUniversity Press, 1994). In addition, certain properties of NMDAreceptors suggest that they may be involved in theinformation-processing in the brain that underlies consciousness itself.

The NMDA receptor has drawn particular interest since it appears to beinvolved in a broad spectrum of CNS disorders. For instance, duringbrain ischemia caused by stroke or traumatic injury, excessive amountsof the excitatory amino acid glutamate are released from damaged oroxygen deprived neurons. This excess glutamate binds to the NMDAreceptors which opens their ligand-gated ion channels; in turn thecalcium influx produces a high level of intracellular calcium whichactivates a biochemical cascade resulting in protein degradation andcell death. This phenomenon, known as excitotoxicity, is also thought tobe responsible for the neurological damage associated with otherdisorders ranging from hypoglycemia and cardiac arrest to epilepsy. Inaddition, there are preliminary reports indicating similar involvementin the chronic neurodegeneration of Huntington's. Parkinson's, andAlzheimer's diseases. Activation of the NMDA receptor has been shown tobe responsible for post-stroke convulsions, and, in certain models ofepilepsy, activation of the NMDA receptor has been shown to be necessaryfor the generation of seizures. Neuropsychiatric involvement of the NMDAreceptor has also been recognized since blockage of the NMDA receptorCa⁺⁺ channel by the animal anesthetic PCP (phencyclidine) produces apsychotic state in humans similar to schizophrenia (reviewed in Johnson,K. and Jones, S., 1990). Further, NMDA receptors have also beenimplicated in certain types of spatial learning.

The NMDA receptor is believed to consist of several protein chainsembedded in the postsynaptic membrane. The first two types of subunitsdiscovered so far form a large extracellular region, which probablycontains most of the allosteric binding sites, several transmembraneregions looped and folded so as to form a pore or channel, which ispermeable to Ca⁺⁺, and a carboxyl terminal region. The opening andclosing of the channel is regulated by the binding of various ligands todomains (allosteric sites) of the protein residing on the extracellularsurface. The binding of the ligands is thought to affect aconformational change in the overall structure of the protein which isultimately reflected in the channel opening, partially opening,partially closing, or closing.

NMDA receptor compounds may exert dual (agonist/antagonist) effect onthe NMDA receptor through the allosteric sites. These compounds aretypically termed “partial agonists”. In the presence of the principalsite ligand, a partial agonist will displace some of the ligand and thusdecrease Ca⁺⁺ flow through the receptor. In the absence of or loweredlevel of the principal site ligand, the partial agonist acts to increaseCa⁺⁺ flow through the receptor channel.

A need continues to exist in the art for novel and more specific/potentcompounds that are capable of binding the glycine binding site of NMDAreceptors, and provide pharmaceutical benefits. In addition, a needcontinues to exist in the medical arts for orally deliverable forms ofsuch compounds.

SUMMARY

Provided herein, at least in part, are compounds that are NMDAmodulators, for example, partial agonists of NMDA. For example,disclosed herein are compounds represented by the formula:

and pharmaceutically acceptable salts, stereoisomers, and N-oxidesthereof, wherein

R_(b) is selected from the group consisting of H, halogen, hydroxyl,cyano or C₁-C₆ alkyl;

R₁ is H or C₁-C₆ alkyl;

R₂ is H or C₁-C₆ alkyl;

R₃ is selected from the group consisting of H, C₁-C₆ alkyl and anitrogen protecting group;

R₄ is H or C₁-C₆ alkyl;

R₅ is X or —C₁-C₆ alkylene-X, wherein X is selected from the groupconsisting of phenyl, a 4- to 6-membered heteroaryl ring having 1, 2, or3 heteroatoms selected from O, S, or N, or a 4- to 6-memberedheterocyclyl ring having 1, 2, or 3 heteroatoms selected from O, S, orN, and wherein X is optionally substituted with

and

R₆ is selected from the group consisting of H, halogen, hydroxyl, cyano,—O—C(O)—C₁-C₆ alkyl, C₁-C₆ alkyl, or C₁-C₆ alkoxy;

or, in other embodiments, the variables set forth in formula (I) aredefined as follows:

R_(b) is selected from the group consisting of H, halogen, hydroxyl,cyano or C₁-C₆ alkyl;

R₁ is H or C₁-C₆ alkyl;

R₂ is H or C₁-C₆ alkyl;

R₃ is selected from the group consisting of H, C₁-C₆ alkyl and anitrogen protecting group;

R₄ is H or C₁-C₆ alkyl;

R₅ is X or —C₁-C₆ alkylene-X, wherein X is selected from the groupconsisting of:

(i) phenyl;

(ii) heteroaryl including from 5 to 6 ring atoms wherein 1, 2, or 3 ofthe ring atoms are independently selected from the group consisting ofN, NH, N(C1-C3 alkyl). O, and S; and

(iii) heterocyclyl including from 3 to 6 ring atoms wherein 1, 2, or 3of the ring atoms are independently selected from the group consistingof N. NH, N(C1-C3 alkyl), O, and S; wherein R₅ is optionally substitutedwith

and

R₆ is selected from the group consisting of H, halogen, hydroxyl, cyano.—O—C(O)—C₁-C₆ alkyl, C₁-C₆ alkyl, or C₁-C₆ alkoxy.

Also disclosed herein are compounds represented by the formula:

and pharmaceutically acceptable salts, stereoisomers, and N-oxidesthereof, whereinR₃ is selected from the group consisting of H, C₁-C₆alkyl and a nitrogenprotecting group; andR₇ is a 4- to 6-membered heteroaryl ring having 1, 2, or 3 heteroatomsselected from O, S, or N, optionally substituted on a free carbon by asubstituent selected from the group consisting of: halogen, C₁-C₆alkyl,hydroxyl, cyano, and phenyl.

Also provided herein are compounds represented by the formula:

and pharmaceutically acceptable salts, stereoisomers, and N-oxidesthereof, whereinR_(b) is selected from the group consisting of H, halogen, hydroxyl,cyano or C₁-C₆ alkyl;R₁ is H or C₁-C₆ alkyl;R₂ is H or C₁-C₆ alkyl;R₃ is selected from the group consisting of H. C₁-C₆ alkyl and anitrogen protecting group;R₄ is H or C₁-C₆ alkyl;X is selected from the group consisting of phenyl, a 4- to 6-memberedheteroaryl ring having 1, 2, or 3 heteroatoms selected from O, S, or N,or a 4- to 6-membered heterocyclyl ring having 1, 2, or 3 heteroatomsselected from O, S, or N, wherein

is present on a free carbon of X; andR₆ is selected from the group consisting of H, halogen, hydroxyl, cyano,—O—C(O)—C₁-C₆ alkyl. C₁-C₆ alkyl, or C₁-C₆ alkoxy.

Also provided herein are pharmaceutically acceptable compositionscomprising a disclosed compound, and a pharmaceutically acceptableexcipient. For example, such compositions may be suitable for oral orintravenous administration to a patient.

In another aspect, a method of treating a condition selected from thegroup consisting of autism, anxiety, depression, bipolar disorder,attention deficit disorder, attention deficit hyperactivity disorder(ADHD), schizophrenia, a psychotic disorder, a psychotic symptom, socialwithdrawal, obsessive-compulsive disorder, phobia, post-traumatic stresssyndrome, a behavior disorder, an impulse control disorder, a substanceabuse disorder, a sleep disorder, a memory disorder, a learningdisorder, urinary incontinence, multiple system atrophy, progressivesupra-nuclear palsy, Friedrich's ataxia. Down's syndrome, fragile Xsyndrome, tuberous sclerosis, olivio-ponto-cerebellar atrophy, cerebralpalsy, drug-induced optic neuritis, ischemic retinopathy, diabeticretinopathy, glaucoma, dementia, AIDS dementia, Alzheimer's disease.Huntington's chorea, spasticity, myoclonus, muscle spasm. Tourette'ssyndrome, epilepsy, cerebral ischemia, stroke, a brain tumor, traumaticbrain injury, cardiac arrest, myelopathy, spinal cord injury, peripheralneuropathy, acute neuropathic pain, and chronic neuropathic, in apatient in need thereof is provided. Such methods may compriseadministering to the patient a pharmaceutically effective amount of adisclosed compound or pharmaceutically acceptable salts, stereoisomers.N-oxides, and hydrates thereof.

In some embodiments, a contemplated method includes treating depression.For example, depression may include one or more of major depressivedisorder, dysthymic disorder, psychotic depression, postpartumdepression, seasonal affective disorder, bipolar disorder, mooddisorder, or depression caused by a chronic medical condition. In otherembodiments, a contemplated method may treat schizophrenia. Suchschizophrenia may be, for example, paranoid type schizophrenia,disorganized type schizophrenia, catatonic type schizophrenia,undifferentiated type schizophrenia, residual type schizophrenia,post-schizophrenic depression, or simple schizophrenia.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the potentiation of [³H]MK-801 binding in the presence ofCompound Y.

DETAILED DESCRIPTION

This disclosure is generally directed to compounds that are capable ofmodulating NMDA. e.g., NMDA antagonists or partial agonists, andcompositions and/or methods of using the disclosed compounds.

Definitions

“Treating” includes any effect, e.g., lessening, reducing, modulating,or eliminating, that results in the improvement of the condition,disease, disorder and the like.

The term “alkenyl” as used herein refers to an unsaturated straight orbranched hydrocarbon having at least one carbon-carbon double bond, suchas a straight or branched group of 2-6 or 3-4 carbon atoms, referred toherein for example as C₂-C₆ alkenyl, and C₃-C₄ alkenyl, respectively.Exemplary alkenyl groups include, but are not limited to, vinyl, allyl,butenyl, pentenyl, etc.

The term “alkoxy” as used herein refers to a straight or branched alkylgroup attached to an oxygen (alkyl-O—). Exemplary alkoxy groups include,but are not limited to, alkoxys of 1-6 or 2-6 carbon atoms, referred toherein as C₁-C₆ alkoxy, and C₂-C₆ alkoxy, respectively. Exemplary alkoxygroups include, but are not limited to methoxy, ethoxy, isopropoxy, etc.

The term “alkenyloxy” used herein refers to a straight or branchedalkenyl group attached to an oxygen (alkenyl-O). Exemplary alkenoxygroups include, but are not limited to, groups with an alkenyl group of3-6 carbon atoms, (also e.g. referred to as C₃-C₆ alkenyloxy). Exemplary“alkenoxy” groups include, but are not limited to allyloxy, butenyloxy,etc.

The term “alkynyloxy” used herein refers to a straight or branchedalkynyl group attached to an oxygen (alkynyl-O)). Exemplary alkynyloxygroups include, but are not limited to, C₃-C₆ alkynyloxy. e.g.,propynyloxy.

The term “alkyl” as used herein refers to a saturated straight orbranched hydrocarbon, such as a straight or branched group of 1-6, 1-4,or 1-3 carbon atoms, referred to herein as C₁-C₆ alkyl. C₁-C₄ alkyl, andC₁-C₃ alkyl, respectively. Exemplary alkyl groups include, but are notlimited to, methyl, ethyl, propyl, isopropyl, 2-methyl-1-propyl,2-methyl-2-propyl, 2-methyl-1-butyl, 3-methyl-1-butyl, 3-methyl-2-butyl,2,2-dimethyl-1-propyl, 2-methyl-1-pentyl, 3-methyl-1-pentyl,4-methyl-1-pentyl, 2-methyl-2-pentyl, 3-methyl-2-pentyl,4-methyl-2-pentyl, 2,2-dimethyl-1-butyl, 3,3-dimethyl-1-butyl,2-ethyl-1-butyl, butyl, isobutyl, t-butyl, pentyl, isopentyl, neopentyl,hexyl, etc. The term “haloalkyl” as used herein refers to a saturatedstraight or branched alkyl groups, in which one or more hydrogen atomsof the alkyl group are replaced with one or more independently selectedhalogens. The term “haloalkyl” encompasses alkyl groups in which all ofhydrogen atoms of the alkyl group are replaced independently selectedhalogens (sometimes referred to as “perhalo” alkyl groups. Exemplaryhaloalkyl groups include, but are not limited to, CH₂F, CH₂CH₂Cl, CF₃,CHFCH₂Cl.

The term “alkynyl” as used herein refers to an unsaturated straight orbranched hydrocarbon having at least one carbon-carbon triple bond, suchas a straight or branched group of 2-6, or 3-6 carbon atoms, referred toherein as C₂-C₆ alkynyl, and C₃-C₆ alkynyl, respectively. Exemplaryalkynyl groups include, but are not limited to, ethynyl, propynyl,butynyl, pentynyl, hexynyl, methylpropynyl, etc.

The term “bridged cycloalkyl”, as used herein, is defined as amonocyclic 4- to 7-membered cycloalkyl group in which two non-adjacentatoms are linked by a CH₂ or CH₂CH₂ group. A “bridged cycloalkyl” may befused to one or more phenyl, partially unsaturated, or saturated rings.Examples of bridged carbocyclic groups include but are not limited tobicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, bicyclo[2.2.2]octene etc.

The term “carbonyl” as used herein refers to the radical —C(O)—. Theterm “cyano” as used herein refers to the radical —CN. The term “nitro”refers to the radical —NO₂. The term “H” refers to hydrogen.

The term “cycloalkoxy” as used herein refers to a cycloalkyl groupattached to an oxygen (cycloalkyl-O—).

The term “cycloalkyl” as used herein refers to a monocyclic saturated orpartially unsatured hydrocarbon group of for example 3-6, or 4-6carbons, referred to herein. e.g., as “C₃₋₆ cycloalkyl” or “C₄₋₆cycloalkyl,” and derived from a cycloalkane. Exemplary cycloalkyl groupsinclude, but are not limited to, cyclohexyl, cyclohexenyl, cyclopentyl,cyclobutyl, cyclopropyl or cyclopentyl.

The terms “halo” or “halogen” as used herein refer to F, Cl, Br, or I.

The terms “heteroaryl” as used herein refers to a monocyclic aromatic4-6 membered ring system containing one or more heteroatoms, for exampleone to three heteroatoms, such as nitrogen, oxygen, and sulfur. Wherepossible, said heteroaryl ring may be linked to the adjacent radicalthough carbon or nitrogen. Examples of heteroaryl rings include but arenot limited to furyl, thienyl, pyrrolyl, thiazolyl, oxazolyl,isothiazolyl, isoxazolyl, imidazolyl, pyrazolyl, triazolyl, oxadiazolyl(e.g., 1,2,4-oxadiazolyl or 1,3,4-oxadiazolyl), pyridyl, andpyrimidinyl.

The terms “heterocyclyl” or “heterocyclic group” are art-recognized andrefer to saturated or partially unsaturated 4- to 7-membered ringstructures, whose ring structures include one to three heteroatoms, suchas nitrogen, oxygen, and sulfur. A heterocycle may be fused to one ormore phenyl, partially unsaturated, or saturated rings. Examples ofheterocyclyl groups include but are not limited pyrrolidinyl,piperidinyl, morpholino, thiomorpholino, and piperazinyl.

The term “heterocyclylalkoxy” as used herein refers to aheterocyclyl-alkyl-O— group.

The term “heterocyclyloxyalkyl” refers to a heterocyclyl-O-alkyl-group.

The term “heterocycloxy” refers to a heterocyclyl-O— group. The term“cycloalkyloxy” refers to a cycloalkyl-O— group.

The term “heteroaryloxy” refers to a heteroaryl-O— group.

The terms “hydroxy” and “hydroxyl” as used herein refers to the radical—OH.

The term “oxo” as used herein refers to the radical ═O.

The term “nitrogen protecting group” or “amino protecting group” isart-recognized and as used herein refers to a chemical moiety that iscovalently linked to a nitrogen atom of an amino (primary or secondary)group and that temporarily blocks the reactivity of the amino groupduring a synthetic step and is selectively removed once the syntheticstep is complete. Nitrogen protecting groups include, for example,9-Fluorenylmethyloxycarbonyl (Fmoc), tert-butoxycarbonyl (Boc),carbobenzyloxycarbonyl (Cbz), p-methoxybenzyloxycarbonyl, acetyl,trifluoroacetyl, benzoyl, phthalimido, benzyl (Bn), p-methoxybenzyl,p-methoxyphenyl, 3,4-dimethoxybenzyl, triphenylmethyl, benzylidene, andp-toluenesulfonyl (Ts). In some embodiments, the nitrogen protectinggroup can have one of the following formulas: —C(O)OR₃₁ or —C(O)R₃₂ asdefined herein. In certain embodiments, R₃₁ is selected from the groupconsisting of: C₁-C₆ alkyl; C₁-C₆ haloalkyl; C₂-C₆ alkenyl; C₂-C₆alkynyl; C₃-C₁₀ cycloalkyl, wherein the C₃-C₁₀ cycloalkyl is optionallysubstituted with from 1-3 independently selected C₁-C₃ alkyl;—CH₂—C₃-C₁₀ cycloalkyl wherein the C₃-C₁₀ cycloalkyl is optionallysubstituted with from 1-3 independently selected C₁-C₃ alkyl;—CH₂-phenyl, wherein the phenyl is optionally substituted with from 1-2substituents independently selected from C₁-C₃ alkyl, C₁-C₃ haloalkyl.C₁-C₃ alkoxy, C₁-C₃ haloalkoxy, nitro, halo, SO₂Me, cyano, and—OC(O)CH₃; and —CH₂-pyridyl. In certain embodiments. R₃₂ is selectedfrom the group consisting of: H; C₁-C₆ alkyl; C₁-C₆ haloalkyl; phenyl,wherein the phenyl is optionally substituted with from 1-2 substituentsindependently selected from C₁-C₃ alkyl, C₁-C₃ haloalkyl. C₁-C₃ alkoxy,C₁-C₃ haloalkoxy, nitro, halo, SO₂Me, cyano, and —OC(O)CH₃: and pyridyl.

As used in the present disclosure, the term “partial NMDA receptoragonist” generally refers to a compound that is capable of binding to aglycine binding site of an NMDA receptor, at low concentrations a NMDAreceptor agonist acts substantially as agonist and at highconcentrations it acts substantially as an antagonist. Theseconcentrations are experimentally determined for each and every “partialagonist.

“Pharmaceutically or pharmacologically acceptable” include molecularentities and compositions that do not produce an adverse, allergic orother untoward reaction when administered to an animal, or a human, asappropriate. For human administration, preparations should meetsterility, pyrogenicity, general safety and purity standards as requiredby FDA Office of Biologics standards.

The term “pharmaceutically acceptable carrier” or “pharmaceuticallyacceptable excipient” as used herein refers to any and all solvents,dispersion media, coatings, isotonic and absorption delaying agents, andthe like, that are compatible with pharmaceutical administration. Theuse of such media and agents for pharmaceutically active substances iswell known in the art. The compositions may also contain other activecompounds providing supplemental, additional, or enhanced therapeuticfunctions.

The term “pharmaceutical composition” as used herein refers to acomposition comprising at least one compound as disclosed hereinformulated together with one or more pharmaceutically acceptablecarriers.

“Individual,” “patient.” or “subject” are used interchangeably andinclude any animal, including mammals, preferably mice, rats, otherrodents, rabbits, dogs, cats, swine, cattle, sheep, horses, or primates,and most preferably humans. The compounds of the invention can beadministered to a mammal, such as a human, but can also be administeredto other mammals such as an animal in need of veterinary treatment,e.g., domestic animals (e.g., dogs, cats, and the like), farm animals(e.g., cows, sheep, pigs, horses, and the like) and laboratory animals(e.g., rats, mice, guinea pigs, and the like). The mammal treated in themethods of the invention is desirably a mammal in which treatment e.g.,of pain or depression is desired. “Modulation” includes antagonism(e.g., inhibition), agonism, partial antagonism and/or partial agonism.

In the present specification, the term “therapeutically effectiveamount” means the amount of the subject compound that will elicit thebiological or medical response of a tissue, system, animal or human thatis being sought by the researcher, veterinarian, medical doctor or otherclinician. The compounds of the invention are administered intherapeutically effective amounts to treat a disease. Alternatively, atherapeutically effective amount of a compound is the quantity requiredto achieve a desired therapeutic and/or prophylactic effect, such as anamount which results in lessening a symptom of depression.

The term “pharmaceutically acceptable salt(s)” as used herein refers tosalts of acidic or basic groups that may be present in compounds used inthe present compositions. Compounds included in the present compositionsthat are basic in nature are capable of forming a wide variety of saltswith various inorganic and organic acids. The acids that may be used toprepare pharmaceutically acceptable acid addition salts of such basiccompounds are those that form non-toxic acid addition salts, i.e., saltscontaining pharmacologically acceptable anions, including but notlimited to malate, oxalate, chloride, bromide, iodide, nitrate, sulfate,bisulfate, phosphate, acid phosphate, isonicotinate, acetate, lactate,salicylate, citrate, tartrate, oleate, tannate, pantothenate,bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate,gluconate, glucaronate, saccharate, formate, benzoate, glutamate,methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonateand pamoate (i.e., 1,1′-methylene-bis-(2-hydroxy-3-naphthoate)) salts.Compounds included in the present compositions that are acidic in natureare capable of forming base salts with various pharmacologicallyacceptable cations. Examples of such salts include alkali metal oralkaline earth metal salts and, particularly, calcium, magnesium,sodium, lithium, zinc, potassium, and iron salts. Compounds included inthe present compositions that include a basic or acidic moiety may alsoform pharmaceutically acceptable salts with various amino acids. Thecompounds of the disclosure may contain both acidic and basic groups;for example, one amino and one carboxylic acid group. In such a case,the compound can exist as an acid addition salt, a zwitterion, or a basesalt.

The compounds of the disclosure may contain one or more chiral centersand/or double bonds and, therefore, exist as stereoisomers, such asgeometric isomers, enantiomers or diastereomers. The term“stereoisomers” when used herein consist of all geometric isomers,enantiomers or diastereomers. These compounds may be designated by thesymbols “R” or “S,” depending on the configuration of substituentsaround the stereogenic carbon atom. The present invention encompassesvarious stereoisomers of these compounds and mixtures thereof.Stereoisomers include enantiomers and diastereomers. Mixtures ofenantiomers or diastereomers may be designated “(±)” in nomenclature,but the skilled artisan will recognize that a structure may denote achiral center implicitly.

The compounds of the disclosure may contain one or more chiral centersand/or double bonds and, therefore, exist as geometric isomers,enantiomers or diastereomers. The enantiomer and diastereomers may bedesignated by the symbols “(+),” “(−).” “R” or “S,” depending on theconfiguration of substituents around the stereogenic carbon atom, butthe skilled artisan will recognize that a structure may denote a chiralcenter implicitly. Geometric isomers, resulting from the arrangement ofsubstituents around a carbon-carbon double bond or arrangement ofsubstituents around a cycloalkyl or heterocyclic ring, can also exist inthe compounds of the present invention. The symbol

denotes a bond that may be a single, double or triple bond as describedherein. Substituents around a carbon-carbon double bond are designatedas being in the “Z” or “E” configuration wherein the terms “Z” and “E”are used in accordance with IUPAC standards. Unless otherwise specified,structures depicting double bonds encompass both the “E” and “Z”isomers. Substituents around a carbon-carbon double bond alternativelycan be referred to as “cis” or “trans,” where “cis” representssubstituents on the same side of the double bond and “trans” representssubstituents on opposite sides of the double bond. The arrangement ofsubstituents around a carbocyclic ring can also be designated as “cis”or “trans.” The term “cis” represents substituents on the same side ofthe plane of the ring and the term “trans” represents substituents onopposite sides of the plane of the ring. Mixtures of compounds whereinthe substituents are disposed on both the same and opposite sides ofplane of the ring are designated “cis/trans.”

The term “stereoisomers” when used herein consist of all geometricisomers, enantiomers or diastereomers. The present invention encompassesvarious stereoisomers of these compounds and mixtures thereof.

Individual enantiomers and diasteriomers of compounds of the presentinvention can be prepared synthetically from commercially availablestarting materials that contain asymmetric or stereogenic centers, or bypreparation of racemic mixtures followed by resolution methods wellknown to those of ordinary skill in the art. These methods of resolutionare exemplified by (1) attachment of a mixture of enantiomers to achiral auxiliary, separation of the resulting mixture of diastereomersby recrystallization or chromatography and liberation of the opticallypure product from the auxiliary, (2) salt formation employing anoptically active resolving agent, (3) direct separation of the mixtureof optical enantiomers on chiral liquid chromatographic columns or (4)kinetic resolution using steroselective chemical or enzymatic reagents.Racemic mixtures can also be resolved into their component enantiomersby well-known methods, such as chiral-phase gas chromatography orcrystallizing the compound in a chiral solvent. Stereoselectivesyntheses, a chemical or enzymatic reaction in which a single reactantforms an unequal mixture of stereoisomers during the creation of a newstereocenter or during the transformation of a pre-existing one, arewell known in the art. Stereoselective syntheses encompass both enantio-and diastereoselective transformations. For examples, see Carreira andKvaerno, Classics in Stereoselective Synthesis, Wiley-VCH: Weinheim,2009.

The compounds disclosed herein can exist in solvated as well asunsolvated forms with pharmaceutically acceptable solvents such aswater, ethanol, and the like, and it is intended that the inventionembrace both solvated and unsolvated forms. In one embodiment, thecompound is amorphous. In one embodiment, the compound is a singlepolymorph. In another embodiment, the compound is a mixture ofpolymorphs. In another embodiment, the compound is in a crystallineform.

The invention also embraces isotopically labeled compounds of theinvention which are identical to those recited herein, except that oneor more atoms are replaced by an atom having an atomic mass or massnumber different from the atomic mass or mass number usually found innature. Examples of isotopes that can be incorporated into compounds ofthe invention include isotopes of hydrogen, carbon, nitrogen, oxygen,phosphorus, fluorine and chlorine, such as ²H, ³H, ¹³C, ¹⁴C, ¹⁵N, ¹⁸O,¹⁷O, ³¹P, ³²P, ³⁵S, ¹⁸F, and ³⁶Cl, respectively. For example, a compoundof the invention may have one or more H atom replaced with deuterium.

Certain isotopically-labeled disclosed compounds (e.g., those labeledwith ³H and ¹⁴C) are useful in compound and/or substrate tissuedistribution assays. Tritiated (i.e., ³H) and carbon-14 (i.e., ¹⁴C)isotopes are particularly preferred for their ease of preparation anddetectability. Further, substitution with heavier isotopes such asdeuterium (i.e., ²H) may afford certain therapeutic advantages resultingfrom greater metabolic stability (e.g., increased in vivo half-life orreduced dosage requirements) and hence may be preferred in somecircumstances. Isotopically labeled compounds of the invention cangenerally be prepared by following procedures analogous to thosedisclosed in the e.g., Examples herein by substituting an isotopicallylabeled reagent for a non-isotopically labeled reagent.

The term “prodrug” refers to compounds that are transformed in vivo toyield a disclosed compound or a pharmaceutically acceptable salt,hydrate or solvate of the compound. The transformation may occur byvarious mechanisms (such as by esterase, amidase, phosphatase, oxidativeand or reductive metabolism) in various locations (such as in theintestinal lumen or upon transit of the intestine, blood or liver).Prodrugs are well known in the art (for example, see Rautio,Kumpulainen, et al, Nature Reviews Drug Discovery 2008, 7, 255). Forexample, if a compound of the invention or a pharmaceutically acceptablesalt, hydrate or solvate of the compound contains a carboxylic acidfunctional group, a prodrug can comprise an ester formed by thereplacement of the hydrogen atom of the acid group with a group such as(C₁-C₈)alkyl, (C₂-C₁₂)alkanoyloxymethyl, 1-(alkanoyloxy)ethyl havingfrom 4 to 9 carbon atoms, 1-methyl-1-(alkanoyloxy)-ethyl having from 5to 10 carbon atoms, alkoxycarbonyloxymethyl having from 3 to 6 carbonatoms, 1-(alkoxycarbonyloxy)ethyl having from 4 to 7 carbon atoms,1-methyl-1-(alkoxycarbonyloxy)ethyl having from 5 to 8 carbon atoms.N-(alkoxycarbonyl)aminomethyl having from 3 to 9 carbon atoms,1-(N-(alkoxycarbonyl)amino)ethyl having from 4 to 10 carbon atoms,3-phthalidyl. 4-crotonolactonyl, gamma-butyrolacton-4-yl,di-N,N—(C₁-C₂)alkylamino(C₂-C₃)alkyl (such as β-dimethylaminoethyl),carbamoyl-(C₁-C₂)alkyl, N,N-di(C₁-C₂)alkylcarbamoyl-(C₁-C₂)alkyl andpiperidino-, pyrrolidino- or morpholino(C₂-C₃)alkyl.

Similarly, if a compound of the invention contains an alcohol functionalgroup, a prodrug can be formed by the replacement of the hydrogen atomof the alcohol group with a group such as (C₁-C₆)alkanoyloxymethyl,1-((C₁-C₆)alkanoyloxy)ethyl, 1-methyl-1-((C₁-C₆)alkanoyloxy)ethyl(C₁-C₆)alkoxycarbonyloxymethyl, N—(C₁-C₆)alkoxycarbonylaminomethyl,succinoyl, (C₁-C₆)alkanoyl. α-amino(C₁-C₄)alkanoyl, arylacyl andα-aminoacyl, or α-aminoacyl-α-aminoacyl, where each α-aminoacyl group isindependently selected from the naturally occurring L-amino acids,P(O)(OH)₂, —P(O)(O(C₁-C₆)alkyl)₂ or glycosyl (the radical resulting fromthe removal of a hydroxyl group of the hemiacetal form of acarbohydrate).

If a compound of the invention incorporates an amine functional group, aprodrug can be formed, for example, by creation of an amide orcarbamate, an N-acyloxyakyl derivative, an (oxodioxolenyl)methylderivative, an N-Mannich base, imine or enamine. In addition, asecondary amine can be metabolically cleaved to generate a bioactiveprimary amine, or a tertiary amine can metabolically cleaved to generatea bioactive primary or secondary amine. For examples, see Simplicio, etal., Molecules 2008, 13, 519 and references therein.

Compounds

Disclosed compounds include those represented by formula I:

and pharmaceutically acceptable salts, stereoisomers, and N-oxidesthereof, wherein

-   -   R_(b) is selected from the group consisting of H, halogen,        hydroxyl, cyano or C₁-C₆ alkyl;    -   R₁ is H or C₁-C₆ alkyl;    -   R₂ is H or C₁-C₆ alkyl;    -   R₃ is selected from the group consisting of H. C₁-C₆ alkyl and a        nitrogen protecting group;    -   R₄ is H or C₁-C₆ alkyl;    -   R₅ is X or —C₁-C₆ alkylene-X, wherein X is selected from the        group consisting of phenyl, a 4- to 6-membered heteroaryl ring        having 1, 2, or 3 heteroatoms selected from O, S, or N, or a 4-        to 6-membered heterocyclyl ring having 1, 2, or 3 heteroatoms        selected from    -   O, S, or N. and wherein R₅ is optionally substituted with

andR₆ is selected from the group consisting of H, halogen, hydroxyl, cyano.—O—C(O)—C₁-C₆ alkyl. C₁-C₆ alkyl, or C₁-C₆ alkoxy; or, in otherembodiments, the variables set forth in formula (I) are defined asfollows:

-   -   R_(b) is selected from the group consisting of H, halogen,        hydroxyl, cyano or C₁-C₆ alkyl (e.g., H);    -   R₁ is H or C₁-C₆ alkyl;    -   R₂ is H or C₁-C₆ alkyl;    -   R₃ is selected from the group consisting of H. C₁-C₆ alkyl and a        nitrogen protecting group;    -   R₄ is H or C₁-C₆ alkyl;    -   R₅ is X or —C₁-C₆ alkylene-X, wherein X is selected from the        group consisting of:    -   (i) phenyl;    -   (ii) heteroaryl including from 5 to 6 ring atoms wherein 1, 2,        or 3 of the ring atoms are independently selected from the group        consisting of N, NH, N(C1-C3 alkyl), O, and S; and    -   (iii) heterocyclyl including from 3 to 6 ring atoms wherein 1,        2, or 3 of the ring atoms are independently selected from the        group consisting of N, NH, N(C1-C3 alkyl), O, and S;        wherein R₅ is optionally substituted with

and

-   -   R₆ is selected from the group consisting of H, halogen,        hydroxyl, cyano, —O—C(O)—C₁-C₆ alkyl, C₁-C₆ alkyl, or C₁-C₆        alkoxy.

In certain embodiments, R₁ is H.

In certain embodiments. R₂ is H.

In certain embodiments, R₃ is H.

In other embodiments, R₃ is a nitrogen protecting group. In certainembodiments. R₃ has formula —C(O)OR₃₁, wherein R₃₁ is selected from thegroup consisting of: C₁-C₆ alkyl; C₁-C₆ haloalkyl; C₂-C₆ alkenyl; C₂-C₆alkynyl; C₃-C₁₀ cycloalkyl, wherein the C₃-C₁₀ cycloalkyl is optionallysubstituted with from 1-3 independently selected C₁-C₃ alkyl;—CH₂—C₃-C₁₀ cycloalkyl wherein the C₃-C₁₀ cycloalkyl is optionallysubstituted with from 1-3 independently selected C₁-C₃ alkyl;—CH₂-phenyl, wherein the phenyl is optionally substituted with from 1-2substituents independently selected from C₁-C₃ alkyl; C₁-C₃ haloalkyl;C₁-C₃ alkoxy; C₁-C₃ haloalkoxy; nitro; halo; SO₂Me, cyano; and—OC(O)CH₃; and —CH₂-pyridyl. In certain embodiments, R₃₁ is C₁-C₆ alkyl(e.g., tert-butyl). In other embodiments, R₃ has formula —C(O)R₃₂,wherein R₃₂ is selected from the group consisting of: H; C₁-C₆ alkyl;C₁-C₆ haloalkyl; phenyl, wherein the phenyl is optionally substitutedwith from 1-2 substituents independently selected from C₁-C₃ alkyl;C₁-C₃ haloalkyl; C₁-C₃ alkoxy; C₁-C₃ haloalkoxy; nitro; halo; SO₂Me,cyano; and —OC(O)CH₃; and pyridyl. In certain embodiments, R₃₂ is C₁-C₆alkyl (e.g., —CH₃ or iso-propyl).

In certain embodiments, R₄ is C₁-C₆ alkyl. In some embodiments. R₄ ismethyl.

In certain embodiments, R_(b) is H.

In certain embodiments, R₅ is X, and wherein X is a 5- to 6-memberedheteroaryl ring selected from the group consisting of azetidine,pyrrolidine, pyrazolidine, pyridine, pyrimidine, isooxazolidine,imidazolidine, oxazolidine, thiazolidine, and isothiazolidine. In otherembodiments, R₅ is

In certain other embodiments, R₅ is X, and wherein X is optionallysubstituted with

(sometimes referred to herein as “—CH(R₄)(R₆)”).

In some embodiments, R₅ is X. In certain embodiments. X is heteroarylincluding from 5 to 6 ring atoms wherein 1, 2, or 3 of the ring atomsare independently selected from the group consisting of N, NH, N(C₁-C₃alkyl), O, and S. For example, X can be selected from the groupconsisting of 1,2,4-oxadiazolyl, 1,3,4-oxadiazolyl, pyridyl, andpyrimidinyl. In certain embodiments, R₅ (here, X as defined anywhereherein) is substituted with —CH(R₄)(R₆). In certain embodiments, R₄ isC₁-C₆ alkyl (e.g., R₄ is methyl). In certain embodiments, R₆ is selectedfrom the group consisting of hydroxyl. —O—C(O)—C₁-C₆ alkyl, and C₁-C₆alkoxy (e.g., R₆ is hydroxyl). Embodiments in which R₅ is X can includeone or more of the following features: R₁ is H or methyl; R₂ is H ormethyl; R₃ is H. —C(O)OR₃₁, or —C(O)R₃₂; when present, R₄ is C₁-C₆ alkyl(e.g., R₄ is methyl), and R₆ is selected from the group consisting ofhydroxyl. —O—C(O)—C₁-C₆ alkyl, and C₁-C₆ alkoxy (e.g., R₆ is hydroxyl);R_(b) is H.

In some embodiments, R₅ is —C₁-C₆ alkylene-X (e.g., —C₁-C₂ alkylene-X or—C₁ alkylene-X). In certain embodiments, X is heteroaryl including from5 to 6 ring atoms wherein 1, 2, or 3 of the ring atoms are independentlyselected from the group consisting of N, NH, N(C₁-C₃ alkyl), O, and S.For example, X can be selected from the group consisting of1,2,4-oxadiazolyl, 1,3,4-oxadiazolyl, pyridyl, and pyrimidinyl. Incertain embodiments. R₅ is substituted on either the X portion or thealkylene chain portion with —CH(R₄)(R₆). In certain of theseembodiments, R₅ is substituted on the alkylene portion with —CH(R₄)(R₆),and R₅ can have, for example, the formula —CH(CHR₄R₆)—X. In certainembodiments, R₄ is C₁-C₆ alkyl (e.g., R₄ is methyl). In certainembodiments, R₆ is selected from the group consisting of hydroxyl,—O—C(O)—C₁-C₆ alkyl, and C₁-C₆ alkoxy (e.g., R₆ is hydroxyl).Embodiments in which R₅ is —C₁-C₆ alkylene-X can include one or more ofthe following features: R₁ is H or methyl; R₂ is H or methyl; R₃ is H,—C(O)OR₃₁, or —C(O)R₃₂; when present. R₄ is C₁-C₆ alkyl (e.g., R₄ ismethyl), and R₆ is selected from the group consisting of hydroxyl,—O—C(O)—C₁-C₆ alkyl, and C₁-C₆ alkoxy (e.g., R₆ is hydroxyl); R_(b) isH.

In certain embodiments. R₆ is hydroxyl.

In some embodiments, a disclosed compound includes those delineated inTable 1 and/or the Examples, e.g., one having the formula:

Disclosed compounds also include compounds represented by formula II:

and pharmaceutically acceptable salts, stereoisomers, and N-oxidesthereof, wherein

-   -   R₃ is selected from the group consisting of H, C₁-C₆alkyl and a        nitrogen protecting group; and    -   R₇ is a 4- to 6-membered heteroaryl ring having 1, 2, or 3        heteroatoms selected from O, S, or N, optionally substituted on        a free carbon by a substituent selected from the group        consisting of: halogen, C₁-C₆alkyl, hydroxyl, cyano, and phenyl.

Also disclosed are compounds represented by formula III:

and pharmaceutically acceptable salts, stereoisomers, and N-oxidesthereof, wherein

-   -   R_(b) is selected from the group consisting of H, halogen,        hydroxyl, cyano or C₁-C₆ alkyl;    -   R₁ is H or C₁-C₆ alkyl;    -   R₂ is H or C₁-C₆ alkyl;    -   R₃ is selected from the group consisting of H. C₁-C₆ alkyl and a        nitrogen protecting group;    -   R₄ is H or C₁-C₆ alkyl;    -   X is selected from the group consisting of phenyl, a 4- to        6-membered heteroaryl ring having 1, 2, or 3 heteroatoms        selected from O, S, or N, or a 4- to 6-membered heterocyclyl        ring having 1, 2, or 3 heteroatoms selected from O, S, or N,        wherein

-   -    is present on a free carbon of X; and    -   R₆ is selected from the group consisting of H, halogen,        hydroxyl, cyano. —O—C(O)—C₁-C₆ alkyl. C₁-C₆ alkyl, or C₁-C₆        alkoxy.

In certain embodiments. R₁ is H.

In certain embodiments, R₂ is H.

In some embodiments, R₃ is H.

In certain embodiments, R₄ is C₁-C₆ alkyl. In other embodiments, R₄ ismethyl.

In some embodiments, R_(b) is H.

In certain embodiments, X is

In some embodiments, R₆ is hydroxyl.

The compounds of the present disclosure and formulations thereof mayhave a plurality of chiral centers. Each chiral center may beindependently R, S, or any mixture of R and S. For example, in someembodiments, a chiral center may have an R:S ratio of between about100:0 and about 50:50, between about 100:0 and about 75:25, betweenabout 100:0 and about 85:15, between about 100:0 and about 90:10,between about 100:0 and about 95:5, between about 100:0 and about 98:2,between about 100:0 and about 99:1, between about 0:100 and 50:50,between about 0:100 and about 25:75, between about 0:100 and about15:85, between about 0:100 and about 10:90, between about 0:100 andabout 5:95, between about 0:100 and about 2:98, between about 0:100 andabout 1:99, between about 75:25 and 25:75, and about 50:50. Formulationsof the disclosed compounds comprising a greater ratio of one or moreisomers (i.e., R and/or S) may possess enhanced therapeuticcharacteristic relative to racemic formulations of a disclosed compoundsor mixture of compounds. In some instances, chemical formulas containthe descriptor “—(R)—” or “—(S)—” that is further attached to solidwedge or dashed wedge. This descriptor is intended to show a methinecarbon (CH) that is attached to three other substituents and has eitherthe indicated R or S configuration (see, e.g., Table 1).

Disclosed compounds may provide for efficient cation channel opening atthe NMDA receptor, e.g. may bind or associate with the glutamate site ofthe NMDA receptor to assist in opening the cation channel. The disclosedcompounds may be used to regulate (turn on or turn off) the NMDAreceptor through action as an agonist.

The compounds as described herein may be glycine site NMDA receptorpartial agonists. A partial agonist as used in this context will beunderstood to mean that at a low concentration, the analog acts as anagonist and at a high concentration, the analog acts as an antagonist.Glycine binding is not inhibited by glutamate or by competitiveinhibitors of glutamate, and also does not bind at the same site asglutamate on the NMDA receptor. A second and separate binding site forglycine exists at the NMDA receptor. The ligand-gated ion channel of theNMDA receptor is, thus, under the control of at least these two distinctallosteric sites. Disclosed compounds may be capable of binding orassociating with the glycine binding site of the NMDA receptor. In someembodiments, disclosed compounds may possess a potency that is 10-foldor greater than the activity of existing NMDA receptor glycine sitepartial agonists.

The disclosed compounds may exhibit a high therapeutic index. Thetherapeutic index, as used herein, refers to the ratio of the dose thatproduces a toxicity in 50% of the population (i.e., TD₅₀) to the minimumeffective dose for 50% of the population (i.e., ED₅₀). Thus, thetherapeutic index=(TD₅₀):(ED₅₀). In some embodiments, a disclosedcompound may have a therapeutic index of at least about 10:1, at leastabout 50:1, at least about 100:1, at least about 200:1, at least about500:1, or at least about 1000:1.

Compositions

In other aspects, formulations and compositions comprising the disclosedcompounds and optionally a pharmaceutically acceptable excipient areprovided. In some embodiments, a contemplated formulation comprises aracemic mixture of one or more of the disclosed compounds.

Contemplated formulations may be prepared in any of a variety of formsfor use. By way of example, and not limitation, the compounds may beprepared in a formulation suitable for oral administration, subcutaneousinjection, or other methods for administering an active agent to ananimal known in the pharmaceutical arts.

Amounts of a disclosed compound as described herein in a formulation mayvary according to factors such as the disease state, age, sex, andweight of the individual. Dosage regimens may be adjusted to provide theoptimum therapeutic response. For example, a single bolus may beadministered, several divided doses may be administered over time or thedose may be proportionally reduced or increased as indicated by theexigencies of the therapeutic situation. It is especially advantageousto formulate parenteral compositions in dosage unit form for ease ofadministration and uniformity of dosage. Dosage unit form as used hereinrefers to physically discrete units suited as unitary dosages for themammalian subjects to be treated; each unit containing a predeterminedquantity of active compound calculated to produce the desiredtherapeutic effect in association with the required pharmaceuticalcarrier.

The specification for the dosage unit forms of the invention aredictated by and directly dependent on (a) the unique characteristics ofthe compound selected and the particular therapeutic effect to beachieved, and (b) the limitations inherent in the art of compoundingsuch an active compound for the treatment of sensitivity in individuals.

Therapeutic compositions typically must be sterile and stable under theconditions of manufacture and storage. The composition can be formulatedas a solution, microemulsion, liposome, or other ordered structuresuitable to high drug concentration. The carrier can be a solvent ordispersion medium containing, for example, water, ethanol, polyol (forexample, glycerol, propylene glycol, and liquid polyethylene glycol, andthe like), and suitable mixtures thereof. The proper fluidity can bemaintained, for example, by the use of a coating such as lecithin, bythe maintenance of the required particle size in the case of dispersionand by the use of surfactants. In many cases, it will be preferable toinclude isotonic agents, for example, sugars, polyalcohols such asmannitol, sorbitol, or sodium chloride in the composition. Prolongedabsorption of the injectable compositions can be brought about byincluding in the composition an agent which delays absorption, forexample, monostearate salts and gelatin.

The compounds can be administered in a time release formulation, forexample in a composition which includes a slow release polymer. Thecompounds can be prepared with carriers that will protect the compoundagainst rapid release, such as a controlled release formulation,including implants and microencapsulated delivery systems.Biodegradable, biocompatible polymers can be used, such as ethylenevinyl acetate, polyanhydrides, polyglycolic acid, collagen,polyorthoesters, polylactic acid and polylactic, polyglycolic copolymers(PLG). Many methods for the preparation of such formulations aregenerally known to those skilled in the art.

Sterile injectable solutions can be prepared by incorporating thecompound in the required amount in an appropriate solvent with one or acombination of ingredients enumerated above, as required, followed byfiltered sterilization. Generally, dispersions are prepared byincorporating the active compound into a sterile vehicle which containsa basic dispersion medium and the required other ingredients from thoseenumerated above. In the case of sterile powders for the preparation ofsterile injectable solutions, the preferred methods of preparation arevacuum drying and freeze-drying which yields a powder of the activeingredient plus any additional desired ingredient from a previouslysterile-filtered solution thereof.

In accordance with an alternative aspect of the invention, a compoundmay be formulated with one or more additional compounds that enhance thesolubility of the compound.

Methods

Methods for treating a condition in a patient in need thereof byadministering a therapeutically effective dose of a compound describedherein are provided. In some embodiments, the condition may be a mentalcondition. For example, a mental illness may be treated. In anotheraspect, a nervous system condition may be treated. For example, acondition that affects the central nervous system, the peripheralnervous system, and/or the eye may be treated. In some embodiments,neurodegenerative diseases may be treated.

In some embodiments, the methods include administering a compound totreat patients suffering from autism, anxiety, depression, bipolardisorder, attention deficit disorder, attention deficit hyperactivitydisorder (ADHD), schizophrenia, a psychotic disorder, a psychoticsymptom, social withdrawal, obsessive-compulsive disorder (OCD), phobia,post-traumatic stress syndrome, a behavior disorder, an impulse controldisorder, a substance abuse disorder (e.g., a withdrawal symptom, opiateaddiction, nicotine addiction, and ethanol addition), a sleep disorder,a memory disorder (e.g., a deficit, loss, or reduced ability to make newmemories), a learning disorder, urinary incontinence, multiple systematrophy, progressive supra-nuclear palsy, Friedrich's ataxia, Down'ssyndrome, fragile X syndrome, tuberous sclerosis,olivio-ponto-cerebellar atrophy, cerebral palsy, drug-induced opticneuritis, ischemic retinopathy, diabetic retinopathy, glaucoma,dementia, AIDS dementia, Alzheimer's disease, Huntington's chorea,spasticity, myoclonus, muscle spasm, Tourette's syndrome, epilepsy,cerebral ischemia, stroke, a brain tumor, traumatic brain injury,cardiac arrest, myelopathy, spinal cord injury, peripheral neuropathy,acute neuropathic pain, and chronic neuropathic pain.

In some embodiments, methods of treating a memory disorder associatedwith aging, schizophrenia, special learning disorders, seizures,post-stroke convulsions, brain ischemia, hypoglycemia, cardiac arrest,epilepsy, migraine, AIDS dementia, Huntington's chorea. Parkinson'sdisease, early stage Alzheimer's disease, and Alzheimer's disease arecontemplated.

In certain embodiments, methods for treating schizophrenia are provided.For example, paranoid type schizophrenia, disorganized typeschizophrenia (i.e., hebephrenic schizophrenia), catatonic typeschizophrenia, undifferentiated type schizophrenia, residual typeschizophrenia, post-schizophrenic depression, and simple schizophreniamay be treated using the methods and compositions contemplated herein.Psychotic disorders such as schizoaffective disorders, delusionaldisorders, brief psychotic disorders, shared psychotic disorders, andpsychotic disorders with delusions or hallucinations may also be treatedusing the compositions contemplated herein.

Paranoid schizophrenia may be characterized where delusions or auditoryhallucinations are present, but thought disorder, disorganized behavior,or affective flattening are not. Delusions may be persecutory and/orgrandiose, but in addition to these, other themes such as jealousy,religiosity, or somatization may also be present. Disorganized typeschizophrenia may be characterized where thought disorder and flataffect are present together. Catatonic type schizophrenia may becharacterized where the patient may be almost immobile or exhibitagitated, purposeless movement. Symptoms can include catatonic stuporand waxy flexibility. Undifferentiated type schizophrenia may becharacterized where psychotic symptoms are present but the criteria forparanoid, disorganized, or catatonic types have not been met. Residualtype schizophrenia may be characterized where positive symptoms arepresent at a low intensity only. Post-schizophrenic depression may becharacterized where a depressive episode arises in the aftermath of aschizophrenic illness where some low-level schizophrenic symptoms maystill be present. Simple schizophrenia may be characterized by insidiousand progressive development of prominent negative symptoms with nohistory of psychotic episodes.

In some embodiments, methods are provided for treating psychoticsymptoms that may be present in other mental disorders, including, butnot limited to, bipolar disorder, borderline personality disorder, drugintoxication, and drug-induced psychosis. In another embodiment, methodsfor treating delusions (e.g., “non-bizarre”) that may be present in, forexample, delusional disorder are provided.

Also provided are methods for treating social withdrawal in conditionsincluding, but not limited to, social anxiety disorder, avoidantpersonality disorder, and schizotypal personality disorder.

In some embodiments, methods are provided for treating neuropathic pain.The neuropathic pain may be acute or chronic. In some cases, theneuropathic pain may be associated with a condition such as herpes, HIV,traumatic nerve injury, stroke, post-ischemia, fibromyalgia, reflexsympathetic dystrophy, complex regional pain syndrome, spinal cordinjury, sciatica, phantom limb pain, diabetic neuropathy, and cancerchemotherapeutic-induced neuropathic pain. Methods for enhancing painrelief and for providing analgesia to a patient are also contemplated.

Further contemplated methods include a method of treating autism and/oran autism spectrum disorder in a patient need thereof, comprisingadministering an effective amount of a compound to the patient. In anembodiment, a method for reducing the symptoms of autism in a patient inneed thereof is contemplated, comprising administering an effectiveamount of a disclosed compound to the patient. For example, uponadministration, the compound may decrease the incidence of one or moresymptoms of autism such as eye contact avoidance, failure to socialize,attention deficit, poor mood, hyperactivity, abnormal sound sensitivity,inappropriate speech, disrupted sleep, and perseveration. Such decreasedincidence may be measured relative to the incidence in the untreatedindividual or an untreated individual(s).

Also provided herein is a method of modulating an autism target geneexpression in a cell comprising contacting a cell with an effectiveamount of a compound described herein. The autism gene expression may befor example, selected from ABAT, APOE. CHRNA4. GABRA5, GFAP, GRIN2A,PDYN, and PENK. In another embodiment, a method of modulating synapticplasticity in a patient suffering from a synaptic plasticity relateddisorder is provided, comprising administering to the patient aneffective amount of a compound.

In another embodiment, a method of treating Alzheimer's disease, ore.g., treatment of memory loss that e.g., accompanies early stageAlzheimer's disease, in a patient in need thereof is provided,comprising administering a compound. Also provided herein is a method ofmodulating an Alzheimer's amyloid protein (e.g., beta amyloid peptide.e.g. the isoform Aβ₁₋₄₂), in-vitro or in-vivo (e.g. in a cell)comprising contacting the protein with an effective amount of a compoundis disclosed. For example, in some embodiments, a compound may block theability of such amyloid protein to inhibit long-term potentiation inhippocampal slices as well as apoptotic neuronal cell death. In someembodiments, a disclosed compound may provide neuroprotective propertiesto a Alzheimer's patient in need thereof, for example, may provide atherapeutic effect on later stage Alzheimer's-associated neuronal celldeath.

In a further embodiment, a method of treating depression comprisingadministering a compound described herein is provided. In someembodiments, the treatment may relieve depression or a symptom ofdepression without affecting behavior or motor coordination and withoutinducing or promoting seizure activity. Exemplary depression conditionsthat are expected to be treated according to this aspect of theinvention include, but are not limited to, major depressive disorder,dysthymic disorder, psychotic depression, postpartum depression,premenstrual syndrome, premenstrual dysphoric disorder, seasonalaffective disorder (SAD), bipolar disorder (or manic depressivedisorder), mood disorder, and depressions caused by chronic medicalconditions such as cancer or chronic pain, chemotherapy, chronic stress,and post traumatic stress disorders. In addition, patients sufferingfrom any form of depression often experience anxiety. Various symptomsassociated with anxiety include fear, panic, heart palpitations,shortness of breath, fatigue, nausea, and headaches among others.Anxiety or any of the symptoms thereof may be treated by administering acompound as described herein.

Also provided herein are methods of treating a condition intreatment-resistant patients, e.g., patients suffering from a mental orcentral nervous system condition that does not, and/or has not,responded to adequate courses of at least one, or at least two, othercompounds or therapeutics. For example, provided herein is a method oftreating depression in a treatment resistant patient, comprising a)optionally identifying the patient as treatment resistant and b)administering an effective dose of a compound to said patient.

In some embodiments, a compound described herein may be used for acutecare of a patient. For example, a compound may be administered to apatient to treat a particular episode (e.g., a severe episode) of acondition contemplated herein.

Also contemplated herein are combination therapies comprising a compoundin combination with one or more other active agents. For example, acompound may be combined with one or more antidepressants, such astricyclic antidepressants. MAO-I's, SSRI's, and double and triple uptakeinhibitors and/or anxiolytic drugs. Exemplary drugs that may be used incombination with a compound include Anafranil, Adapin. Aventyl, Elavil,Norpramin. Pamelor, Pertofrane, Sinequan. Surmontil, Tofranil, Vivactil.Parnate, Nardil. Marplan, Celexa, Lexapro, Luvox, Paxil. Prozac. Zoloft,Wellbutrin. Effexor. Remeron. Cymbalta, Desyrel (trazodone), andLudiomill. In another example, a compound may be combined with anantipsychotic medication. Non-limiting examples of antipsychoticsinclude butyrophenones, phenothiazines, thioxanthenes, clozapine,olanzapine, risperidone, quetiapine, ziprasidone, amisulpride,asenapine, paliperidone, iloperidone, zotepine, sertindole, lurasidone,and aripiprazole. It should be understood that combinations of acompound and one or more of the above therapeutics may be used fortreatment of any suitable condition and are not limited to use asantidepressants or antipsychotics.

EXAMPLES

The following examples are provided for illustrative purposes only, andare not intended to limit the scope of the disclosure.

Table 1 below shows some exemplary compounds of the disclosure andprovides physiochemical characteristics of the compounds.

TABLE 1 Molecular Weight Compound Structure (Da) cLogP tPSA Compound X

252 −1.14 91.5 Compound Y

262 −0.57 78.35 Compound Z

248 −0.15 78.35 C-16

252.2697 −1.1382 91.49 C-4

262.3076 −0.57075 78.35 C-15

352.3856 0.144989 109 C-20

276.3342 −0.160256 78.35 C-36

276.3342 −0.160256 78.35 C-33

290.3608 0.250239 78.35 C-10

252.2697 −1.83479 91.49 C-9

352.3856 −0.551597 109 C-3

362.4234 0.672806 95.86 C-23

366.4121 −0.135022 109 C-19

376.45 1.0833 95.86 C-26

366.4121 −0.135022 109 C-29

376.45 1.0833 95.86 C-32

390.4766 1.4938 95.86 C-11

322.3596 −0.981326 99.77 C-5

332.3975 0.335914 86.63 C-37

346.424 0.746408 86.63

Example 1—Synthesis of Compound X

Synthesis of 2-((tert-butoxycarbonyl) amino)-3-hydroxybutanoic Acid (A)

To a stirring solution of 2-amino-3-hydroxybutanoic acid (SM2) (10 g,83.9 mmol) in 1,4-dioxane/water (100 mL, 1:1)) was added NaHCO₃ (21.1 g,0.25 mol) followed by Boc-anhydride (21.9 mL, 0.101 mol) at 0° C. Thereaction mixture was stirred at RT for 16 h. After consumption of thestarting material (by TLC), the reaction mixture was diluted with waterand washed with EtOAc. The aqueous layer was acidified using citric acidsolution (pH˜3-4) and then extracted with CH₂Cl₂ (2×150 mL). Theseparated organic extracts were dried over anhydrous Na₂SO₄, filteredand concentrated under vacuum to afford A (15 g, crude). This materialwas directly used for the next step without further purification.

Synthesis of 3-(tert-butoxycarbonyl)-2, 2,5-trimethyloxazolidine-4-carboxylic Acid (B)

To a stirring solution of A (15 g, 59.28 mmol) in THF (150 mL) was addedPPTS (1.47 g, 5.92 mmol) followed by 2,2-dimethoxy propane (21.79 mL,0.17 mol) at 0° C. under N₂ atmosphere. The reaction mixture was stirredat RT for 16 h. The reaction mixture was again heated to reflux for 6 h.The reaction mixture was diluted with aqueous NaHCO₃ solution and washedwith EtOAc. The aqueous layer was acidified using citric acid solution(pH˜2) and extracted with CH₂Cl₂ (2×100 mL). The organic layer waswashed with brine, dried over anhydrous Na₂SO₄ and concentrated undervacuum to afford B (18 g, crude).

¹H-NMR: (400 MHz, DMSO-d₆): δ 13.25 (br s, 1H), 4.11-4.05 (m, 1H), 3.79(d, 1H), 1.50 (s, 3H), 1.67 (s, 3H), 1.45 (s, 9H), 1.29 (d, 3H).

Synthesis of tert-butyl 4-carbamoyl-2, 2,5-trimethyloxazolidine-3-carboxylate (C)

To a stirring solution of B (18 g, 69.4 mmol) in CH₂Cl₂ (180 mL) wasadded HOBt (14.16 g, 0.104 mol). EDCI.HCl (19.88 g, 0.104 mol) followedby NH₄Cl (5.56 g, 0.104 mol) and DIPEA (31.9 mL, 0.173 mol) at 0° C. Thereaction mixture was stirred at RT for 16 h. After consumption of thestarting material (by TLC), the reaction mixture was washed with aqueouscitric acid, NaHCO₃ followed by brine. The organic layer was dried overanhydrous Na₂SO₄ and concentrated under reduced pressure. The resultingcrude material was purified by silica gel column chromatography elutingwith 2% MeOH/CH₂Cl₂ to afford C (13 g, 72.5%).

¹H-NMR: (400 MHz, DMSO-d₆): δ 7.51 (br s, 1H), 7.14 (br s, 1H),3.97-3.95 (m, 1H), 3.71 (d, 1H), 1.51 (d, 6H), 1.34 (s, 9H), 1.24 (d,3H).

LCMS (ESI): 159.1 [(M⁺+1)-Boc]

Synthesis of (Z)-tert-butyl 4-(((dimethylamino) methylene)carbamoyl)-2,2,5-trimethyloxazolidine-3-carboxylate (D)

A solution of C (13 g, 50.3 mmol) in DMF.DMA (130 mL) was stirred atreflux temperature for 3 h under N₂ atmosphere. After consumption of thestarting material (by TLC), the reaction mixture was concentrated underreduced pressure to afford D (15.7 g, crude). This crude material wasdirectly taken for the next step without further purification.

Synthesis of tert-butyl 2,2,5-trimethyl-4-(1,2,4-oxadiazol-5-yl)oxazolidine-3-carboxylate (E)

To a stirring solution of D (15.7 g, 50.09 mmol) in ethanol (157 mL) wasadded hydroxylamine hydrochloride (6.96 g, 0.10 mol) under N₂atmosphere. The reaction mixture was heated to reflux and stirred for 2h. After consumption of the starting material (by TLC), acetic acid(28.6 mL, 0.50 mol) was added to the reaction mixture and then refluxedfor 16 h. Solvents from the reaction mixture were evaporated undervacuum. The resulting crude material was purified by silica gel columnchromatography eluting with 10% EtOAc/Hexane to afford E (4.5 g, 32%).

¹H-NMR: (400 MHz, DMSO-d₆): δ 6.35 (s, 2H), 4.61 (d, 1H), 4.22-4.15 (m,1H), 1.55 (s, 6H), 1.37 (s, 2H), 1.25 (d, 3H), 1.21 (s, 6H).

LCMS (ESI): 284 [M⁺+1]

Mass (m/z): 283 [M⁺]

Synthesis of 1-amino-1-(1, 2, 4-oxadiazol-5-yl) propan-2-ol (Int-F)

To a stirring solution of E (5 g, 17.6 mmol) in water (25 mL) was addedtrifluoroacetic acid (25 mL). The reaction mixture was stirred at RT for5 h. After consumption of the starting material (by TLC), the reactionmixture was concentrated under vacuum. The residue was dissolved inwater and neutralized with aqueous NaHCO₃. The solvents from thereaction mixture were evaporated under vacuum and extracted with 5%MeOH/CH₂C₂ (4×150 mL). The organic layer was concentrated under reducedpressure to afford Int-F (2.5 g, crude).

¹H-NMR: (400 MHz, D₂O): δ 8.84 (s, 1H), 4.05 (d, 1H), 3.98-3.95 (m, 1H),3.67 (s, 1H), 3.58 (d, 1H), 1.15 (d, 3H), 1.12 (d, 3H).

LCMS (ESI): 144.1 [M⁺+1]

Synthesis of methyl pyrrolidine-2-carboxylate (1)

To a stirring solution of pyrrolidine-2-carboxylic acid (SM1) (100 g,0.87 mol) in methanol (800 mL) was added thionyl chloride (76.9 mL, 1.04mol) slowly drop wise at 0° C. The reaction mixture was heated to refluxfor 12 h. After consumption of the starting material (by TLC), thereaction was concentrated under vacuum. Obtained residue was washed withn-Hexane and distilled off the solvent to afford 1 (143.9 g, HCl salt).

¹H-NMR: (400 MHz, CDCl₃) (Rotamers): δ 3.89 (s, 3H), 3.68-3.62 (m, 2H),3.59-3.47 (m, 2H), 2.49-2.37 (m, 1H), 2.27-2.05 (m, 3H).

LCMS (ESI): 166 [M⁺+1]

Synthesis of 1-tert-butyl 2-methyl pyrrolidine-1,2-dicarboxylate (2)

To a stirring solution of 1 (35 g, 0.22 mol) in CH₂Cl₂ (175 mL) wereadded Et₃N (90 mL, 0.65 mol) followed by Boc-anhydride (56.9 mL, 0.26mol) at 0° C. The reaction mixture was stirred at RT for 16 h. Afterconsumption of the starting material (by TLC), the reaction was dilutedwith water (100 mL) and extracted with CH₂Cl₂ (2×100 mL). The organiclayer was washed with water, brine, dried over Na₂SO₄ and concentrated.Obtained crude material was purified by silica gel column chromatographyeluting with 30% EtOAc/Hexane to afford 2 (41 g, 95%).

¹H-NMR: (400 MHz, CDCl₃) (Rotamers): δ 4.25-4.21 (m, 1H), 3.75 (s, 3H),3.57-3.26 (m, 2H), 2.29-2.10 (m, 1H), 1.99-1.75 (m, 3H), 1.45 (s, 9H).

LCMS (ESI): 130 [(M⁺+1)-Boc]

Synthesis of 1-tert-butyl 2-methyl 2-((benzyloxy) methyl) pyrrolidine-1,2-dicarboxylate (3)

To a stirring solution of 2 (100 g, 0.43 mol) in THF (800 mL) was addedLiHMDS (873 mL, 0.87 mol) at −78° C. and stirred for 1 h. To thisBOM-chloride (93.2 mL, 0.65 mol) was added drop wise at −78° C. andstirred for 2 h at −20° C. After consumption of the starting material(by TLC), the reaction was quenched with NH₄Cl at 0° C. The separatedorganic layer was washed with water, dried over Na₂SO₄ and concentratedto afford 3 (180 g. crude). This material was directly taken for thenext step without further purification.

LCMS (ESI): 250 [(M⁺+1)-Boc]

Synthesis of 1-tert-butyl 2-methyl 2-(hydroxymethyl)pyrrolidine-1,2-dicarboxylate (4)

To a stirring solution of 3 (74 g, 0.21 mol) in methanol (740 mL) wasadded 10% Pd/C (50% wet, 14.8 g) under N₂ atmosphere and stirred for 6 hunder H₂ atmosphere (balloon pressure). The reaction mixture wasfiltered through celite pad and concentrated under reduced pressure toafford 4 (45 g, 82%) as crude.

Synthesis of 1-tert-butyl 2-methyl 2-formylpyrrolidine-1,2-dicarboxylate (5)

To a stirring solution of 4 (10 g, 38.5 mmol) in CH₂Cl₂ (100 mL) wasadded Dess-martin periodinane (19.6 g, 46.27 mmol) at 0° C. under N₂atmosphere and stirred for 3 h. After consumption of the startingmaterial (determined by TLC), the reaction was quenched with aqueousNaHCO₃ solution and extracted with CH₂Cl₂ (2×100 mL). The organic layerwas dried over anhydrous Na₂SO₄ and concentrated under vacuum. Theresulting crude material was purified by column chromatography elutingwith 10% EtOAc/Hexane to afford 5 (7 g, 70.5%).

Synthesis of 1-tert-butyl 2-methyl 2-((((1S, 2R)-2-hydroxy-1-(1, 2,4-oxadiazol-5-yl) propyl) amino) methyl) pyrrolidine-1, 2-dicarboxylate(6)

To a stirring solution of 5 (3 g, 11.6 mmol) in MeOH (30 mL) was addedsodium acetate (1.91 g, 23.3 mmol) followed by Int-F (3.6 g, 13.9 mmol).The reaction mixture was heated to reflux for 1 h. The reaction mixturewas slowly cooled to RT-0° C. to this sodium cyanoborohydride (1.465 g,23.3 mmol) was added, and stirring was continued for another 6 h at RT.After consumption of the starting material (determined by TCL), methanolfrom the reaction was evaporated under reduced pressure and the residuewas diluted with water and extracted with EtOAc (2×50 mL). The separatedorganic layer was dried over anhydrous Na₂SO₄ and concentrated underreduced pressure. The resulting crude material was purified by silicagel column chromatography eluting with 40% EtOAc/Hexane to afford 6 (2.5g, 56%).

LCMS m/z: 385 [M⁺+1]

Synthesis of tert-butyl 2-((1S, 2R)-2-hydroxy-1-(1, 2, 4-oxadiazol-5-yl)propyl)-1-oxo-2, 5-diazaspiro[3.4] octane-5-carboxylate (7)

To a stirring solution of 6 (1.5 g, 3.90 mmol) in THF (30 mL) was cooledto 0° C. and added t-BuMgCl (1M in THF, 15.6 mL, 15.6 mmol) and stirredfor 15 min. After consumption of the starting material (by TLC), thereaction mixture was quenched with aqueous NH₄Cl solution and dilutedwith water. Aqueous layer was extracted with EtOAc (2×25 mL). Theseparated organic layer was dried over anhydrous Na₂SO₄ and concentratedunder reduced pressure. The resulting crude material was purified bysilica gel column chromatography eluting with 150% EtOAc/CH₂Cl₂ toafford 7 (0.15 g, 11%).

¹H-NMR: (400 MHz, DMSO-d6): δ 9.02 (s, 1H), 5.15 (s, 1H), 4.37-4.32 (m,1H), 3.95 (d, 1H), 3.66-3.60 (m, 1H), 3.36-3.30 (m, 1H), 2.29-2.09 (m,2H), 1.87-1.82 (m, 2H), 1.55 (s, 9H), 1.27 (d, 3H).

LCMS (ESI) m/z: 351 [M⁺−1]

UPLC Purity: 96%

Synthesis of 2-((1S, 2R)-2-hydroxy-1-(1, 2, 4-oxadiazol-5-yl) propyl)-2,5-diazaspiro [3.4] octan-1-one (Compound X)

To a stirring solution of 7 (0.4 g, 1.13 mmol) in CH₂Cl₂ (4 mL) wasadded TFA (0.43 mL) at 0° C. and stirred at RT for 30 min. The reactionmixture was concentrated under vacuum. The resulting crude material waspurified by prep-HPLC to afford Compound X (65 mg) as TFA salt.

¹H-NMR: (400 MHz, DMSO-d6): δ 9.89 (br s, 1H), 9.08 (s, 1H), 5.46 (d,1H), 5.31 (s, 1H), 4.37-4.35 (m, 1H), 3.99 (d, 1H), 3.81 (d, 1H),3.42-3.35 (m, 2H), 2.35-2.18 (m, 2H), 2.10-2.03 (m, 2H), 1.24 (d, 3H).

LCMS (ESI) m/z: 253.4 [M⁺×1]

Prep-HPLC Purity: 95%

Example 2—Synthesis of Compound Y

Synthesis of (S)-methyl pyrrolidine-2-carboxylate (A)

To a stirring solution of L-proline (SM1) (100 g, 0.87 mol) in methanol(800 mL) was slowly added thionyl chloride (76.9 mL, 1.04 mol) at 0° C.The reaction mixture was slowly warmed to RT and heated to reflux for 12h. After consumption of the starting material (by TLC), the reaction wasconcentrated under reduced pressure to afford A (143.9 g, HCl salt).

¹H-NMR: (400 MHz, CDCl₃): δ 3.89 (s, 3H), 3.68-3.62 (m, 2H), 3.59-3.47(m, 2H), 2.49-2.37 (m, 1H), 2.27-2.05 (m, 3H).

LCMS (m/z): 166 [M⁺+1]

Synthesis of (S)-1-tert-butyl 2-methyl pyrrolidine-1,2-dicarboxylate (B)

To a stirring solution of A (35 g, 0.22 mol) in CH₂Cl₂ (175 mL) wereadded Et₃N (90 mL, 0.65 mol) followed by Boc-anhydride (56.9 mL, 0.26mol) at 0° C. The reaction mixture was stirred at RT for 16 h. Afterconsumption of the starting material (by TLC), the reaction was dilutedwith water (100 mL) and extracted with CH₂Cl₂ (2×100 mL). The organiclayer was washed with water, brine, dried over Na₂SO₄ and concentrated.The resulting crude material was purified by silica gel columnchromatography eluting with 30% EtOAc/Hexane to afford B (41 g, 95%).

¹H-NMR: (400 MHz, CDCl₃): δ 4.25-4.21 (m, 1H), 3.75 (s, 3H), 3.57-3.26(m, 2H), 2.29-2.10 (m, 1H), 1.99-1.75 (m, 3H), 1.45 (s, 9H).

LCMS (m/z): 130 [(M⁺+1)-Boc]

Synthesis of 1-tert-butyl 2-methyl 2-((benzyloxy) methyl) pyrrolidine-1,2-dicarboxylate (C)

To a stirring solution of B (100 g, 0.43 mol) in THF (800 mL) was addedLiHMDS (873 mL, 0.87 mol) at −78° C. and stirred for 1 h. To thisBOM-chloride (93.2 mL, 0.65 mol) was added drop wise at −78° C. andstirred for 2 h at −20° C. After consumption of the starting material(by TLC), the reaction was quenched with sat.NH₄Cl solution at 0° C. Theseparated organic layer was washed with water, dried over Na₂SO₄ andconcentrated to afford C (180 g, crude). This material was directlytaken for the next step without further purification.

LCMS (m/z): 250 [(M⁺+1)-Boc]

Synthesis of 2-((benzyloxy) methyl)-1-(tert-butoxycarbonyl)pyrrolidine-2-carboxylic Acid (D)

To a stirring solution of C (100 g, 0.28 mol) in methanol (200 mL) wasadded 2N NaOH solution (300 mL) at RT. The reaction mixture was heatedto reflux for 4 h. After consumption of the starting material (by TLC),the solvent from the reaction was evaporated under reduced pressure anddiluted with EtOAc (100 mL). The aqueous layer was acidified usingcitric acid solution and extracted with CH₂Cl₂ (2×250 mL). The separatedorganic layer was washed with water, dried over Na₂SO₄ and concentratedto afford D (60 g, 63%).

¹H-NMR: (400 MHz. CDCl₃): δ 7.37-7.32 (m, 5H), 4.61 (s, 2H), 4.05-3.88(m, 2H), 3.65-3.42 (m, 2H), 2.54-2.46 (m, 2H), 1.95 (br s, 2H), 1.57 (s,9H).

LCMS (m/z): 334 [M⁺−1]

Synthesis of1-(tert-butoxycarbonyl)-2-(hydroxymethyl)pyrrolidine-2-carboxylic Acid(Int-E)

To a stirring solution of D (10 g, 29.81 mmol) in methanol (300 mL) wasadded 50% wet 10% Pd/C (5 g) at RT and stirred for 24 h under H₂atmosphere (balloon pressure). After consumption of the startingmaterial (by TLC), the reaction mixture was filtered through a pad ofcelite and the pad was washed with methanol. The filtrate wasconcentrated under reduced pressure to afford Int-E (6 g, 82%).

¹H-NMR: (400 MHz, DMSO-d₆): δ 12.55 (br m, 1H), 3.99 (d, 1H), 3.88 (d,1H), 7.65-7.60 (m, 1H), 3.51-3.45 (m, 1H), 3.39-3.34 (m, 1H), 2.32-2.14(m, 1H), 1.98-1.69 (m, 3H), 1.39 (s, 9H).

Synthesis of 1-(pyrimidin-2-yl)propan-1-one (2)

To a stirring solution of 1 (15 g, 0.14 mol) in THF (150 mL) was addedethylmagnesium bromide (1M in THF) (171.4 mL, 0.17 mol) at 0° C. undernitrogen atmosphere slowly over a period of 15 min. After being stirredfor 30 min, the reaction mixture was quenched with aqueous NH₄Clsolution and the aqueous layer was extracted with EtOAc. The separatedorganic layer was dried over anhydrous Na₂SO₄, filtered and concentratedunder reduced pressure to obtain crude product, which was purified bysilica gel column chromatography eluting with 50% EtOAc/hexane to afford2 (8.5 g, 44%).

¹H-NMR: (400 MHz, CDCl₃): δ 8.95 (d, 2H), 7.47 (t, 1H), 3.26 (q, 2H),1.25 (t, 3H).

Synthesis of(Z)-2-(1-((tert-butyldimethylsilyl)oxy)prop-1-en-1-yl)pyrimidine (3)

To a stirring solution of 2 (8.5 g, 62.5 mmol) in THF (170 mL) was addedLiHMDS (1M in THF) (93.7 mL, 93.7 mmol) and stirred for 1 h. To this wasadded TES-Cl (12.5 mL, 75 mmol) at 0° C. under N₂ atmosphere and furtherstirred for 1 h. After consumption of the starting material (by TLC),the reaction was quenched with aqueous NH₄Cl solution and extracted withEtOAc. The separated organic layer was dried over anhydrous Na₂SO₄,filtered and concentrated under reduced pressure to afford 3 (10 g,64%). The resulting crude material was directly used for the next stepwithout further purification.

Synthesis of 2-bromo-1-(pyrimidin-2-yl) propan-1-one (4)

To a stirring solution of 3 (10 g, 0.04 mol) in THF: H₂O (100 mL, 4:1)was added NBS (5.6 g, 0.05 mol) at RT and stirred for 1 h. Afterconsumption of the starting material (by TLC), the reaction mixture wasconcentrated under reduced pressure and obtained residue was dilutedwith EtOAc and washed with water. The separated organic layer was driedover anhydrous Na₂SO₄ filtered and concentrated under reduced pressureto obtain crude product, which was purified by silica gel columnchromatography eluting with 30% EtOAc/hexane to afford 4 (6.5 g, 75.5%).

¹H-NMR: (400 MHz, CDCl₃): δ 8.97 (d, 2H), 7.52 (t, 1H), 5.92 (q, 1H),2.79 (s, 1H), 1.99 (d, 3H).

Synthesis of 2-hydroxy-1-(pyrimidin-2-yl) propan-1-one (5)

To a stirring solution of 4 (6.5 g, 0.03 mol) in MeOH (65 mL) was addedsodium formate (10.28 g, 0.15 mol) and heated to reflux for 8 h. Afterconsumption of the starting material (by TLC) the reaction mixture wascooled to RT and solvent was removed under reduced pressure. Theresulting crude material was purified by silica gel columnchromatography eluting with 2% MeOH/CH₂Cl₂ to afford 5 (3.5 g, 76%).

¹H-NMR: (400 MHz. CDCl₃): δ 9.01 (d, 2H), 7.52 (t, 1H), 5.92 (q, 1H),3.73 (br s, 1H), 3.41 (s, 1H), 2.78 (s, 2H), 1.55 (d, 3H).

Synthesis of2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl)propan-1-one (6)

To a stirring solution of 5 (3.5 g, 23 mmol) in CH₂Cl₂ (70 mL) was addedimidazole (3.91 g, 57 mmol) followed by DMAP (281 mg, 2.30 mmol) at RT.The reaction mixture was cooled to 0° C., tow which TBS-Cl (5.18 g, 0.03mol) was added and stirred at RT for 4 h. After consumption of thestarting material (by TLC), the reaction mixture was diluted with CH₂Cl₂and washed with water. The separated organic layer was dried overanhydrous Na₂SO₄, filtered and concentrated under reduced pressure toobtain a crude product, which was purified by silica gel columnchromatography eluting with 20% EtOAc/hexane to afford 6 (3 g, 49%).

¹H-NMR: (400 MHz. CDCl₃: δ 8.93 (d, 2H), 7.45 (t, 1H), 5.59 (q, 1H),1.52 (d, 3H), 0.89 (s, 9H), 0.09 (s, 6H).

Synthesis of2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl)propan-1-amine (7)

To a stirring solution of 6 (3.0 g, 11.27 mmol) in MeOH (60 mL) wasadded sodium acetate (1.84 g, 22.55 mmol) followed by ammonium carbonate(8.85 g, 56.3 mmol) and acetic acid (0.64 mL, 11.27 mmol) at RT. Thereaction mixture was heated to reflux for 1 h. added NaCNBH₃ (1.41 g,22.5 mmol) and continued reflux for another 6 h. After consumption ofthe starting material (by TLC) the reaction mixture was cooled to RT andvolatiles were evaporated. Obtained residue was diluted with water andextracted with EtOAc. The separated organic layer was dried overanhydrous Na₂SO₄ filtered and concentrated under reduced pressure to getcrude product, which was purified by silica gel column chromatographyeluting with 2% MeOH/CH₂Cl₂ to afford 7 (2 g, 66.4%).

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.91 (d, 1H), 8.85 (d, 1H), 7.48 (t, 1H),5.71 (br m, 2H), 4.24 (t, 1H), 4.05 (d, 1H), 1.29 (d, 1H), 1.12 (d, 2H),0.74 (s, 9H), 0.03 (s, 3H), 0.02 (s, 2H), 0.01 (s, 1H).

LCMS (m/z): 268 [M⁺+1]

Synthesis of tert-butyl2-((2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl)propyl)carbamoyl)-2-(hydroxymethyl)pyrrolidine-1-carboxylate(8)

To a stirring solution of Int-E (2 g, 8.16 mmol) in CH₂Cl₂ (60 mL) wasadded compound 7 (2.39 g, 8.97 mmol), EDCI. HCl (2.33 g, 12.2 mmol)followed by HOBt (1.66 g, 12.24 mmol) and DIPEA (4.5 mL, 24.4 mmol) at0° C. The reaction mixture was warmed to RT and stirred for 16 h. Afterconsumption of the starting material (by TLC) the reaction mixture wasdiluted with CH₂Cl₂. The separated organic layer was washed with aqueousNaHCO₃ solution followed by aqueous NH₄Cl. The organic layer was driedover anhydrous Na₂SO₄, filtered and concentrated under reduced pressureto get crude product, which was purified by silica gel columnchromatography eluting with 70% EtOAc/hexane to afford 8 (2.3 g, 57.5%).

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.91 (d, 1H), 8.68 (d, 2H), 7.41 (br s,1H), 5.74 (br t, 1H), 5.07-4.89 (m, 1H), 4.15-4.10 (m, 1H), 3.97-3.92(m, 1H), 3.45-3.41 (m, 1H), 1.79-1.74 (m, 2H), 1.43-1.39 (m, 4H),1.29-1.21 (m, 6H), 1.12 (d, 5H), 0.71 (s, 9H), 0.12 (t, 1H), 0.09 (s,2H), 0.08 (s, 1H), 0.04 (s, 2H).

LCMS (m/z): 495.5 [M⁺+1]

Mass: 495.5 [M⁺+1]

Synthesis of tert-butyl2-(2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl)propyl)-1-oxo-2,5-diazaspiro[3.4]octane-5-carboxylate(9)

To a stirring solution of 8 (2.3 g, 4.65 mmol) in THF (23 mL) was addedTPP (1.34 g, 5.12 mmol) followed by DTAD (1.6 g, 6.98 mmol) at 0° C. Thereaction mixture was warmed to RT and stirred for 16 h. Afterconsumption of the starting material (by TLC), the reaction mixture wasconcentrated under reduced pressure to get crude product which waspurified by silica gel column chromatography eluting with 25%EtOAc/hexane to afford 9 (1.2 g, 54.2%).

LCMS (m/z): 477.4 [M⁺+1]

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.82 (d, 2H), 7.49 (t, 1H), 4.72 (d, 1H),4.31 (q, 1H), 3.62 (br s, 2H), 3.25-3.19 (m, 1H), 2.24-2.05 (m, 2H),1.85-1.81 (m, 2H), 1.42 (br s, 1H), 1.25 (t, 3H), 0.92 (s, 8H), 0.75 (s,9H), 0.02 (s, 3H).

Synthesis of tert-butyl2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-1-oxo-2,5-diazaspiro[3.4]octane-5-carboxylate(10)

To a stirring solution of 9 (1.0 g, 2.10 mmol) in THF (20 mL) was addedTBAF (1M in THF) (6.3 mL, 6.30 mmol) at 0° C. under N₂ atmosphere andstirred for 1 h. After consumption of the starting material (by TLC) thereaction mixture was quenched with ice water and extracted with EtOAc.The separated organic layer was dried over anhydrous Na₂SO₄, filteredand concentrated under reduced pressure to obtain crude product, whichwas purified by silica gel column chromatography eluting with 5%MeOH/CH₂Cl₂ to afford 10 (0.35 g, 46%).

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.81 (d, 2H), 7.49 (t, 1H), 4.81 (d, 1H),4.65 (d, 1H), 4.25-4.20 (m, 1H), 3.64-3.51 (m, 2H), 3.34 (s, 1H),3.25-3.20 (m, 1H), 2.25-2.20 (m, 2H), 1.87-1.82 (m, 2H), 1.19 (d, 3H),0.97 (s, 9H).

LCMS (m/z): 363.3 [M⁺+1]

Synthesis of2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-2,5-diazaspiro[3.4]octan-1-one(Compound Y)

To a stirring solution of 10 (0.3 g, 0.82 mmol) in CH₂Cl₂ (6 mL) wasadded molecular sieves (0.3 g) followed by BF₃-etherate (0.31 mL, 2.48mmol) at 0° C. and stirred at RT for 1 h. After consumption of thestarting material (by TLC), the reaction mixture was filtered andobtained residue was dissolved in MeOH and washed with CH₂Cl₂. Thevolatiles were evaporated under reduced pressure to obtain crudeproduct, which was purified by silica gel column chromatography elutingwith 8% MeOH/CH₂Cl₂ to afford Compound Y (0.12 g, 55%).

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.81 (d, 2H), 7.49 (t, 1H), 4.81 (d, 1H),4.65 (d, 1H), 4.25-4.20 (m, 1H), 3.64-3.51 (m, 2H), 3.12-3.01 (m, 2H),2.15-2.10 (m, 2H), 1.87-1.82 (m, 2H), 1.19 (d, 3H).

LCMS (m/z): 263.1 [M⁺+1]

Preparation of Key Intermediates, Schemes I-1 to I-7:

Synthesis of 1-(pyrimidin-2-yl) propan-1-one, A

Referring to Scheme I-1, to a stirring solution ofpyrimidine-2-carbonitrile (25 g, 238 mmol) in THF (180 mL) was addedethyl magnesium bromide (1M in THF, 37.9 g, 285 mmol) at 0° C. andstirred for 2 h at RT. After completion of starting material (by TLC),the reaction mixture was diluted with saturated ammonium chloridesolution and EtOAc (150 mL). The separated organic layer was washed withbrine solution (2×100 mL). The extracted organic layer was dried overanhydrous Na₂SO₄ filtered and concentrated under reduced pressure toafford crude material which was purified by column chromatographyeluting 30% EtOAc/hexane to afford A (18.3 g, 57%) as an off-whitesolid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.00 (d, J=5.2 Hz, 2H), 7.70 (t, J=4.8 Hz,1H), 3.20-3.15 (m, 2H), 1.09 (t, J=7.2 Hz, 3H).

LCMS m/z: 137 [M⁺+1].

Synthesis of (Z)-2-(1-((triethylsilyl) oxy) prop-1-en-1-yl) pyrimidine(B)

To a stirring solution of A (14 g, 103 mmol) in dry THF (140 mL) wasadded LiHMDS (1M in THF, 206 mL, 206 mmol) slowly at 0° C. and stirredfor 30 min. After added chloro triethylsilane (24.8 g, 165 mmol) in THF(50 mL) dropwise at 0° C. and stirred 1 h. After completion of startingmaterial (by TLC) the reaction mixture was diluted with saturatedammonium chloride solution (100 mL) and EtOAc (150 mL). The separatedorganic layer was washed with brine solution (2×100 mL). The organiclayer was dried over anhydrous Na₂SO₄, filtered and concentrated underreduced pressure to afford crude material which was purified by columnchromatography eluting 20% EtOAc/hexane to afford B (23 g, 89.4%) asyellow thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.75 (d, J=4.8 Hz, 2H), 7.32 (t, J=4.8 Hz,1H), 6.36-6.31 (m, 1H), 1.77 (d, J=7.2 Hz, 3H), 0.95-0.87 (m, 9H),0.71-0.65 (m, 6H).

Synthesis of 2-bromo-1-(pyrimidin-2-yl) propan-1-one (C)

To a stirring solution of B (23 g, 92 mmol) in THF/H₂O (184 mL/46 mL)were added N-bromosuccinamide (18 g, 101 mmol) slowly at 0° C. andstirred for 3 h at RT. After completion of starting material (by TLC),the reaction mixture was diluted with H₂O and EtOAc (100 ml/150 mL). Theseparated organic layer was washed with hypo solution (2×100 mL)followed by brine solution (2×100 mL). The organic layer was dried overanhydrous Na₂SO₄ filtered and concentrated under reduced pressure toafford crude material which was purified by column chromatographyeluting 15% EtOAc/hexane to afford C (11.5 g, 58%) as yellow thicksyrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.06 (d, J=4.8 Hz, 2H), 7.75 (t, J=4.8 Hz,1H), 5.97-5.92 (m, 1H), 1.83 (d, J=6.4 Hz, 3H).

Synthesis of 2-hydroxy-1-(pyrimidin-2-yl) propan-1-one (D)

To a stirring solution of C (11.5 g, 53.4 mmol) in MeOH (80 mL) wasadded sodium formate (14.5 g, 214 mmol) and stirred the reaction mass at80° C. for 6 h. After completion of reaction (by TLC), the reactionmixture was evaporated under reduced pressure to give crude product,which was purified by column chromatography eluting 50% EtOAc/n-hexaneto afford D (5.0 g, 61.6%) as colorless liquid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.73 (d, J=5.2 Hz, 2H), 7.55 (t, J=4.8 Hz,1H), 5.28-5.26 (m, 1H), 1.24 (d, J=6.4 Hz, 1H), 0.99 (d, J=6.4 Hz, 3H)

Synthesis of (2-((tert-butyldimethylsilyl) oxy)-1-(pyrimidin-2-yl)propan-1-one (E)

To a stirring solution of D (5 g, 32.8 mmol) in DCM (50 mL) were addedimidazole (5.5 g, 82.2 mmol). DMAP (800 mg, 0.65 mmol) at 0° C. andstirred for 10 min. After added TBDMS-Cl (7.38 g, 49.2 mmol) at 0° C.and stirred at RT for 12 h. After completion of starting material (byTLC), diluted the reaction mass with H₂O (50 mL). The separated organiclayer was washed with brine solution (2×50 mL). The organic layer wasdried over anhydrous Na₂SO₄ filtered and concentrated under reducedpressure to afford crude material which was purified by columnchromatography eluting 20% EtOAc/hexane to afford E (6 g, 68.8%) as anoff-white solid.

¹H-NMR: (400 MHz. CDCl₃): δ 9.00 (d, J=5.2 Hz, 2H), 7.71 (t, J=4.8 Hz,1H), 5.47-5.42 (m, 1H), 1.35 (d, J=6.8 Hz, 3H), 0.79 (s, 9H), 0.05 (s,6H).

Synthesis of (2-((tert-butyldimethylsilyl) oxy)-1-(pyrimidin-2-yl)propan-1-amine (F)

To a stirring solution of E (6 g, 22.5 mmol) in MeOH (50 mL) were addedsodium acetate (3.69 g, 45.1 mmol), ammonium carbonate (21 g, 135 mmol),AcOH (1.28 mL, 22.5 mmol) at RT and stirred the reaction mixture at 90°C. for 2 h. The temperature of the reaction was cooled to RT and addedsodium cyanoborohydride (2.84 g, 45.1 mmol) slowly and stirred at 90° C.for 6 h. After completion of reaction (by TLC) evaporated MeOH underreduced pressure. The crude residue was diluted with water (20 mL) andextracted with DCM (2×50 mL). The organic layer was dried over anhydrousNa₂SO₄, filtered and concentrated under reduced pressure to afford crudematerial was purified by column chromatography eluting 5% MeOH/DCM toafford F (4.2 g, 69.6%) as semi solid.

¹H-NMR: (400 MHz. CDCl₃): δ 8.83 (d, J=4.8 Hz, 2H), 7.40 (t, J=5.2 Hz,1H), 4.13 (t, J=6.4 Hz, 2H), 3.90 (d, J=6.4 Hz, 2H), 1.12 (d, J=6.4 Hz,3H), 0.70 (s, 9H), 0.02 (s, 6H).

Synthesis of 1-tert-butyl 2-methyl 2-((benzyloxy) methyl) pyrrolidine-1,2-dicarboxylate (G)

To a stirring solution of t-BOC proline methyl ester (25 g, 109 mmol) inTHF (250 mL) was added LiHMDS (240 mL, 240 mmol) at −20 OC and stirredfor 2 h. To this BOM-chloride (23 mL, 163 mmol) was added drop wise at−30° C. and stirred for 2 h. After consumption of the starting material(by TLC), the reaction was quenched with aqueous NH₄Cl solution (100 mL)and extracted with EtOAc (2×200 mL). The combined organic layer waswashed with water (2×150 mL) followed by brine solution (2×100 mL). Theorganic layer was dried over Na₂SO₄ and concentrated to obtain crudecompound which was purified by column chromatography by eluting 10%EtOAC/n-hexane to afford compound G (30 g, 79%) as thick syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 7.36-7.22 (m, 5H), 4.59-4.48 (m, 2H),4.02-3.88 (m, 1H), 3.63 (s, 3H), 3.49-3.35 (m, 2H), 3.34-3.30 (m, 1H),2.31-2.23 (m, 1H), 2.04-1.89 (m, 2H), 1.82-1.78 (m, 1H).

LCMS (m/z): 249.4 [(M⁺+1)-Boc].

Synthesis of 2-((benzyloxy) methyl)-1-(tert-butoxycarbonyl)pyrrolidine-2-carboxylic Acid (H)

To a stirring solution of compound G (30 g, 86 mmol) in methanol (70 mL)was added NaOH solution (6.88 g in 70 mL H₂O) at RT. The reactionmixture was heated to 70° C. for 16 h. After consumption of the startingmaterial (by TLC), the solvent from the reaction was evaporated underreduced pressure and diluted with EtOAc (2×200 mL). The separatedaqueous layer was acidified using citric acid solution (pH˜3) andextracted with EtOAc (2×250 mL). The combined organic layer was driedover Na₂SO₄ and concentrated to afford crude which was triturated withn-hexane to obtain compound H (25 g, 86.8%) as off-white solid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 12.35 (br s, 1H), 7.37-7.29 (m, 5H),4.56-4.48 (m, 2H), 4.06-4.00 (m, 1H), 3.92-3.89 (m, 1H), 3.66-3.45 (m,1H), 3.37-3.28 (m, 1H), 2.31-2.20 (m, 1H), 2.05-1.97 (m, 1H), 1.87-1.75(m, 2H), 1.38 (s, 9H).

LCMS (m/z): 335.3 [M⁺+1].

Synthesis of 1-(tert-butoxycarbonyl)-2-(hydroxymethyl)pyrrolidine-2-carboxylic Acid (I)

To a stirring solution of compound H (25 g, 74 mmol) in methanol (150mL) was added 50% wet 10% Pd/C (7 g) at RT and stirred for 10 h under H₂atmosphere. After consumption of the starting material (by TLC), thereaction mixture was filtered through a pad of celite and the pad waswashed with methanol (100 mL). Obtained filtrate was concentrated underreduced pressure to afford compound I (15 g, 82.8%) as white solid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 4.66 (br s, 1H), 3.96-3.83 (m, 1H),3.63-3.59 (m, 1H), 3.49-3.41 (m, 1H), 3.34-3.25 (m, 1H), 2.30-2.17 (m,1H), 1.95-1.72 (m, 3H), 1.38 (s, 9H).

Synthesis of (2S, 3R)-methyl 2-(((benzyloxy) carbonyl)amino)-3-((tert-butyldimethylsilyl) oxy) butanoate (K)

Referring to Scheme I-3, to a stirring solution of J (50 g, 187 mmol) inDMF (400 mL) were added DIPEA (86 mL, 468 mmol) TBDMS-Cl (33.66 mL, 224mmol) at 0° C. and stirred at RT for 12 h. After completion of startingmaterial (by TLC) diluted the reaction mass with EtOAc (500 ml). Theseparated organic layer was washed with (2×200 mL) of Water followed bybrine solution (2×100 mL). The organic layer was dried over anhydrousNa₂SO₄ filtered and concentrated under reduced pressure to afford crudematerial was purified by column chromatography eluting 10% EtOAc/hexaneto afford K (50 g, 70.1%) as colorless syrup.

¹H-NMR: (400 MHz, CDCl₃): δ 7.39-7.32 (m, 5H), 5.43 (d, J=9.6 Hz, 1H),5.14 (s, 2H), 4.45-4.43 (m, 1H), 4.29-4.26 (m, 1H), 3.72 (s, 3H), 1.21(d, J=6.0 Hz, 3H), 0.83 (s, 9H), 0.09 (s, 6H).

LCMS m/z: 382.2 [M⁺+1].

Synthesis of benzyl ((2S, 3R)-3-((tert-butyldimethylsilyl)oxy)-1-hydrazinyl-1-oxobutan-2-yl) carbamate (L)

A solution of K (50 g, 131 mmol) in EtOH (400 mL) was added hydrazinehydrate (32.8 g, 656 mmol), at RT and after stirred at 90° C. for 24 h.After completion of starting material (by TLC), evaporated ethanol underreduced pressure. The crude residue was diluted with water (100 mL) andEtOAc (500 mL). After the separated organic layer was washed with (2×100mL) of water followed by brine solution (1×100 mL). Dried over anhydrousNa₂SO₄, filtered and concentrated under reduced pressure. Obtained crudematerial was purified by column chromatography by eluting with 20%EtOAc/hexane to afford L (25 g, 50%) as colorless thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.10 (s, 1H), 7.36-7.30 (m, 5H), 6.83 (d,J=9.6 Hz, 1H), 5.02 (s, 2H), 4.19 (s, 2H), 4.05-4.02 (m, 1H), 3.97-3.93(m, 1H), 1.05 (d, J=6.0 Hz, 3H), 0.81 (s, 9H), 0.01 (s, 6H).

Synthesis of benzyl ((1S, 2R)-2-((tert-butyldimethylsilyl) oxy)-1-(1, 3,4-oxadiazol-2-yl) propyl) carbamate (M)

A solution of L (25 g, 65.6 mmol) in triethyl orthoformate (250 mL) wasadded p-TSA (catalytic, 250 mg) at RT and after stirred at 80° C. for 3h. After completion of starting material (by TLC), evaporated triethylorthoformate under reduced pressure. The crude residue was purified bycolumn chromatography eluting 10% EtOAc/hexane to afford M (15 g, 58%)as thick syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.22 (s, 1H), 7.85 (d, J=9.5 Hz, 1H),7.36-7.31 (m, 5H), 5.05 (s, 2H), 4.96-4.93 (m, 1H), 4.25 (t, J=6.0 Hz,1H), 1.23 (d, J=6.0 Hz, 3H), 0.80 (s, 9H), 0.10 (s, 6H).

LCMS m/z: 392.4 [M⁺+1].

Synthesis of (1S, 2R)-2-((tert-butyldimethylsilyl) oxy)-1-(1, 3,4-oxadiazol-2-yl) propan-1-amine (N)

To a stirring solution of M (15 g, 38.3 mmol) in methanol (200 mL) wasadded 50% wet 10% Pd/C (5 g) and stirred under H₂ atmosphere (balloonpressure) for 4 h at RT. The reaction mixture was filtered through a padof celite and triturated with methanol (100 mL). The filtrate wasconcentrated under reduced pressure to afford N (10 g, crude) as thicksyrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.15 (s, 1H), 4.11 (t, J=5.0 Hz, 1H), 4.03(d, J=2.0 Hz, 1H), 2.05 (br s, 2H), 1.17 (d, J=6.0 Hz, 3H), 0.76 (s,9H), 0.02 (s, 6H).

LCMS m/z: 258.3 [M⁺+1].

Synthesis of 3-(tert-butoxycarbonyl)-2, 2,5-trimethyloxazolidine-4-carboxylic Acid (O)

To a stirring solution of N—BOC threonine (15 g, 59.28 mmol) in THF (150mL) was added PPTS (1.47 g, 5.92 mmol) followed by 2,2-dimethoxy propane(21.79 mL, 0.17 mol) at 0° C. under N₂ atmosphere. The reaction mixturewas stirred at RT for 16 h. The reaction mixture was again heated toreflux for 6 h. The reaction mixture was diluted with aqueous NaHCO₃solution and washed with EtOAc (lx 100 mL). Aqueous layer was acidifiedusing citric acid solution (pH˜2) and extracted with CH₂C₂ (2×100 mL).The organic layer was washed with brine, dried over anhydrous Na₂SO₄ andconcentrated under vacuum to afford compound O (18 g, crude).

¹H-NMR: (400 MHz, DMSO-d₆): δ13.25 (br s, 1H), 4.11-4.05 (m, 1H), 3.79(d, 1H), 1.50 (s, 3H), 1.67 (s, 3H), 1.45 (s, 9H), 1.29 (d, 3H).

Synthesis of tert-butyl 4-carbamoyl-2, 2,5-trimethyloxazolidine-3-carboxylate (P)

To a stirring solution of compound O (18 g, 69.4 mmol) in CH₂Cl₂ (180mL) was added HOBt (14.16 g, 0.104 mol). EDCI.HCl (19.88 g, 0.104 mol)followed by NH₄Cl (5.56 g, 0.104 mol) and DIPEA (31.9 mL, 0.173 mol) at0° C. The reaction mixture was stirred at RT for 16 h. After consumptionof the starting material (by TLC) the reaction mixture was washed withaqueous citric acid, NaHCO₃ followed by brine. Organic layer was driedover anhydrous Na₂SO₄ and concentrated under reduced pressure to givecrude; which was purified by silica gel column chromatography elutingwith 2% MeOH/CH₂Cl₂ to afford compound P (13 g, 72.5%).

¹H-NMR: (400 MHz, DMSO-d₆): δ 7.51 (br s, 1H), 7.14 (br s, 1H),3.97-3.95 (m, 1H), 3.71 (d, 1H), 1.51 (d, 6H), 1.34 (s, 9H), 1.24 (d,3H).

LCMS (ESI): 159.1 [(M⁺+1)-Boc]

Synthesis of (Z)-tert-butyl 4-(((dimethylamino) methylene) carbamoyl)-2,2, 5-trimethyloxazolidine-3-carboxylate (Q)

A solution of compound P (13 g, 50.3 mmol) in DMF.DMA (130 mL) wasstirred at reflux temperature for 3 h under N₂ atmosphere. Afterconsumption of the starting material (by TLC) the reaction mixture wasconcentrated under reduced pressure to afford compound Q (15.7 g,crude). This crude material was directly taken for the next step withoutfurther purification.

Synthesis of tert-butyl 2, 2, 5-trimethyl-4-(1, 2, 4-oxadiazol-5-yl)oxazolidine-3-carboxylate (R)

To a stirring solution of compound Q (15.7 g, 50.09 mmol) in ethanol(157 mL) was added hydroxylamine hydrochloride (6.96 g, 0.10 mol) underN₂ atmosphere. The reaction mixture was heated to reflux and stirred for2 h. After consumption of the starting material (by TLC), acetic acid(28.6 mL, 0.50 mol) was added to the reaction mixture and then refluxedfor 16 h. The solvents from the reaction mixture was evaporated undervacuum to give crude; which was purified by silica gel columnchromatography eluting with 10% EtOAc/Hexane to afford compound R (4.5g, 32%).

¹H-NMR: (400 MHz, DMSO-d₆): δ 6.35 (s, 2H), 4.61 (d, 1H), 4.22-4.15 (m,1H), 1.55 (s, 6H), 1.37 (s, 2H), 1.25 (d, 3H), 1.21 (s, 6H).

LCMS (ESI): 284 [M⁺+1]

Mass (m/z): 283 [M⁺]

Synthesis of 1-amino-1-(1, 2, 4-oxadiazol-5-yl) propan-2-ol (S)

To a stirring solution of compound R (5 g, 17.6 mmol) in water (25 mL)was added trifluoroacetic acid (25 mL). The reaction mixture was stirredat RT for 5 h. After consumption of the starting material (by TLC), thereaction mixture was concentrated under vacuum. The residue wasdissolved in water and neutralized with aqueous NaHCO₃. The solvent fromthe reaction mixture was evaporated under vacuum and extracted with 5%MeOH/CH₂Cl₂ (3×100 mL). The organic layer was concentrated under reducedpressure to afford compound S (2.5 g. crude).

¹H-NMR: (400 MHz, D₂O): δ 8.84 (s, 1H), 4.05 (d, 1H), 3.98-3.95 (m, 1H),3.67 (s, 1H), 3.58 (d, 1H), 1.15 (d, 3H), 1.12 (d, 3H).

LCMS (ESI): 144.1 [M⁺+1]

Synthesis of ethyl 2-((tert-butoxycarbonyl) amino)-5-oxohexanoate (T)

To a stirring solution of 1-tert-butyl 2-ethyl5-oxopyrrolidine-1,2-dicarboxylate (12 g, 46.6 mmol) in THF (120 mL)under inert atmosphere was added MeMgBr (3M in ether) (20.2 mL, 60.6mmol) at 0° C. and stirred for 2 h. After consumption of the startingmaterial (by TLC), the reaction mixture was quenched with aqueous NH₄Clsolution and the aqueous layer was extracted with EtOAc (2×200 mL). Thecombined organic extracts were dried over anhydrous Na₂SO₄ andconcentrated under reduced pressure. The crude residue obtained waspurified by silica gel column chromatography eluting with 20%EtOAc/Hexane to afford compound T (10 g, 79%).

¹H-NMR: (400 MHz. CDCl₃): δ 5.14 (br s, 1H), 4.23 (q, 2H), 2.62-2.47 (m,2H), 2.17 (s, 4H), 1.91-1.82 (m, 1H), 1.45 (s, 10H), 1.26 (t, 3H).

Synthesis of ethyl 5-methylpyrrolidine-2-carboxylate (V)

To a stirring solution of compound T (10 g, 36.7 mmol) in CH₂Cl₂ (50 mL)was added TFA (14.89 mL, 194.6 mmol) at 0° C. After being stirred for 2h at RT, the reaction mixture was concentrated under reduced pressure toget compound U. Obtained material was dissolved in ethanol (0.100 mL)and 10% Pd/C (50% wet, 3 g) under N₂ atmosphere. The reaction mixturewas stirred under H₂ atmosphere (balloon pressure) for 16 h. Thereaction mixture was filtered through a pad of celite and filtrate wasconcentrated under reduced pressure to afford compound V (15 g, crude).This material was directly taken for the next step without furtherpurification.

¹H-NMR: (500 MHz, DMSO-d₆): 4.4 (m, 1H), 4.2 (m, 2H), 3.6 (m, 1H), 2.3(m, 1H), 2.1 (m, 2H), 1.6 (m, 1H), 1.3 (d, 3H), 1.2 (t, 3H).

LCMS m/z: 158.1 [M⁺+1].

Synthesis of 1-tert-butyl 2-ethyl 5-methylpyrrolidine-1, 2-dicarboxylate(W)

To a stirring solution of compound V (30 g, 191 mmol) in CH₂Cl₂ (150 mL)was added DMAP (23.3 g, 191 mmol) followed by Et₃N (79.8 mL, 573 mmol)and Boc-anhydride (104 mL, 477 mmol) at 0° C. The reaction mixture wasstirred at RT for 16 h. The reaction mixture was diluted with CH₂Cl₂ (50mL) and washed with water (2×150 mL) followed by brine. The separatedorganic layer was dried over anhydrous Na₂SO₄ and concentrated undervacuum. Obtained crude material was purified by column chromatographyeluting with 6% EtOAc/Hexane to afford compound W (12 g, 82%) as paleyellow liquid.

¹H-NMR: (500 MHz, DMSO-d₆): δ4.13-3.86 (m, 4H), 2.15 (d, J=3.5 Hz, 1H),1.99-1.82 (m, 2H), 1.52 (t, J=4.5 Hz, 1H), 1.38 (s, 9H), 1.24 (t, J=5.5Hz, 3H), 1.16 (d, J=6.5 Hz, 3H)

LCMS (m/z): 258 [(M⁺+1).

Synthesis of 1-tert-butyl 2-ethyl 2-((benzyloxy)methyl)-5-methylpyrrolidine-1, 2-dicarboxylate (X)

To a stirring solution of compound W (8.0 g, 31.12 mmol) in THF (70 mL)was added LiHMDS (59 mL, 41.72 mmol) at −78° C. and stirred for 2 h. Tothis BOM-chloride (6.56 mL, 41.72 mmol) was added drop wise and stirredfor 2 h at −30° C. After consumption of the starting material (by TLC),the reaction was quenched with aqueous NH₄Cl solution (20 mL) andextracted with DCM (30 mL). The separated organic layer was dried overNa₂SO₄ and concentrated to afford crude material was purified by columnchromatography eluting with 10% EtOAc/Hexane to afford compound X (11 g,94.2%) as pale yellow liquid.

¹H-NMR: (500 MHz, DMSO-d₆): δ 7.33-7.25 (m, 5H), 4.38 (d, J=10.5 Hz,2H), 4.08-3.98 (m, 1H), 3.88 (d, J=9.5 Hz, 2H), 2.20-2.08 (m, 2H), 1.38(s, 9H), 1.37-1.29 (m, 4H), 1.19 (t, J=7.5 Hz, 3H), 1.14-1.10 (m, 3H).

LCMS (m/z): 378 (M⁺+1).

Synthesis of 2-((benzyloxy)methyl)-1-(tert-butoxycarbonyl)-5-methylpyrrolidine-2-carboxylic Acid(Y)

To a stirring solution of compound X (11 g, 29.17 mmol) in CH₃OH/THF (22mL/20 mL) were added 2N NaOH solution (33 mL) at RT. The reactionmixture was heated to 65° C. for 8 h. After consumption of the startingmaterial (by TLC), the solvent from the reaction was evaporated underreduced pressure and diluted with EtOAc (50 mL). The aqueous layer wasacidified using citric acid solution and extracted with CH₂Cl₂ (2×100mL). The separated organic layer was washed with water (1×50 mL), driedover Na₂SO₄ and concentrated to afford compound Y (8 g, 80%).

¹H-NMR: (400 MHz, DMSO-d₆): δ 12.58 (s, 1H), 7.34-7.28 (m, 5H),4.54-4.47 (m, 2H), 4.05-3.87 (m, 2H), 3.70-3.62 (m, 1H), 2.28-2.08 (m,3H), 1.46-1.37 (m, 1H), 1.28 (s, 9H).

LCMS (m/z): 350 [M⁺+1].

Synthesis of1-(tert-butoxycarbonyl)-2-(hydroxymethyl)-5-methylpyrrolidine-2-carboxylicAcid (Z)

To a stirring solution of compound Y (8 g, 1.45 mmol) in methanol (40mL) was added 50% wet Pd/C (4 g) under N₂ atmosphere. The reactionmixture was stirred under H₂ atmosphere (balloon pressure) at RT for 16h. After consumption of the starting material (by TLC), the reactionmixture was filtered through a pad of celite and the pad was washed withmethanol. Obtained filtrate was concentrated under reduced pressure toafford crude compound which was triturated with n-pentane to obtainedcompound Z (4.5 g, 75.2%) as white solid.

¹H-NMR: (500 MHz, DMSO-d₆): δ12.37 (br s, 1H), 4.61 (br s, 1H),3.95-3.85 (m, 3H), 2.18-2.06 (m, 3H), 1.44-1.41 (m, 1H), 1.38 (s, 9H),1.09 (d, J=6.0 Hz, 3H); LCMS (ESI): 260 [M⁺+1]

Synthesis of 1-(tert-butyl) 2-methyl 2-(1-hydroxyethyl) pyrrolidine-1,2-dicarboxylate (AA)

To a stirring solution of N—BOC proline methyl ester (30 g, 131 mmol) inTHF (100 mL) was added LiHMDS (0.323 mL, 327 mmol) at −20° C. andstirred for 30 min. To this acetaldehyde (12.3 mL, 196 mmol) was addeddrop wise at −20 OC and stirred for 2 h. After consumption of thestarting material (by TLC), the reaction was quenched with aqueous NH₄Clsolution (100 mL) and extracted with EtOAc (2×200 mL). The combinedorganic layer was washed with brine solution (2×150 mL). The organiclayer was dried over Na₂SO₄ and concentrated to obtain crude compoundwhich was purified by column chromatography by eluting 10%EtOAc/n-hexane to afford compound AA (30 g, 83.8%) as thick syrup.

¹H-NMR: (400 MHz, CDCl₃): δ 5.82 (d, J=10.0 Hz, 1H), 4.71-4.11 (m, 1H),3.76 (s, 3H), 3.66-3.57 (m, 1H), 3.45-3.36 (m, 1H), 2.51-2.46 (m, 1H),2.33-2.24 (m, 1H), 2.04-1.97 (m, 1H), 1.95-1.83 (m, 1H), 1.49 (s, 9H),1.20-1.11 (m, 3H).

LCMS (m/z): 274.3 [M⁺+1].

Synthesis of 1-(tert-butoxycarbonyl)-2-(1-hydroxyethyl)pyrrolidine-2-carboxylic Acid (BB)

To a stirring solution of compound AA (30 g, 109 mmol) in methanol (50mL) was added NaOH solution (8.72 g in 10 mL H₂O, 218 mmol) at RT. Thereaction mixture was heated to 70° C. for 12 h. After consumption of thestarting material (by TLC), the solvent from the reaction was evaporatedunder reduced pressure and diluted with DCM (200 mL). The separatedaqueous layer was acidified using citric acid solution (pH˜3) andextracted with DCM (2×250 mL). The combined organic layer was washedwith brine solution (0.1×100 mL). The organic layer was dried overNa₂SO₄ and concentrated to afford compound BB (9.5 g, 33.6%) as brownsolid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 4.53-4.45 (m, 1H), 3.59-3.48 (m, 1H),3.25-3.19 (m, 1H), 2.33-2.16 (m, 1H), 1.90-1.78 (m, 3H), 1.38 (s, 9H),0.95 (d, J=6.4 Hz, 3H).

LCMS (m/z): 258.2 [M⁻−1].

Synthesis of 1-tert-butyl 2-ethyl2-(1-hydroxyethyl)-5-methylpyrrolidine-1, 2-dicarboxylate (CC)

To a stirring solution of compound W (18.0 g, 70 mmol) in THF (200 mL)was added LiHMDS (84 mL, 84 mmol) drop wise at −20 OC and stirred for 30min. To this acetaldehyde (4.2 mL, 77 mmol) was added drop wise andstirred for 45 min at −20 OC. After consumption of the starting material(by TLC), the reaction was quenched with aqueous NH₄Cl solution (100 mL)and extracted with EtOAc (2×150 mL). The separated organic layer wasdried over Na₂SO₄ and concentrated to afford crude material was purifiedby column chromatography eluting with 30% EtOAc/Hexane to affordcompound CC (15 g, 71.4%) as colorless syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 7.33-7.25 (m, 5H), 4.38 (d, J=10.5 Hz,2H), 4.08-3.98 (m, 1H), 3.88 (d, J=9.5 Hz, 2H), 2.20-2.08 (m, 2H), 1.38(s, 9H), 1.37-1.29 (m, 4H), 1.19 (t, J=7.5 Hz, 3H), 1.14-1.10 (m, 3H);

LCMS (m/z): 378 (M⁺+1)

Synthesis of1-(tert-butoxycarbonyl)-2-(1-hydroxyethyl)-5-methylpyrrolidine-2-carboxylicAcid (DD)

To a stirring solution of compound CC (15 g, 49 mmol) in CH₃OH/THF (10mL/40 mL) were added NaOH (3.98 g, 99 mmol) in water (10 mL) at RT. Thereaction mixture was heated to 70° C. for 16 h. After consumption of thestarting material (by TLC), the solvent from the reaction was evaporatedunder reduced pressure and acidified by using citric acid (pH˜4). Theaqueous layer was extracted with EtOAc (2×200 mL). The combined organiclayer was dried over Na₂SO₄ and concentrated to obtained crude compound,which was purified by column chromatography eluting 40% EtOAc to affordcompound DD (4 g, 29.4%) as brown syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 12.15 (br s, 2H), 4.54-4.50 (m, 1H),4.03-4.02 (m, 1H), 2.17-1.77 (m, 3H), 1.41 (s, 9H), 1.39-1.09 (m, 3H),0.99-0.94 (m, 3H).

LCMS (m/z): 272.4 [M⁻−1].

Synthesis of tert-butyl2-((2-4(tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl)propyl)carbamoyl)-2-(hydroxymethyl) pyrrolidine-1-carboxylate (C-1)

Referring to Scheme 1, to a stirring solution of I (2 g, 8.16 mmol) inCH₂Cl₂ (60 mL) was added intermediate F (2.39 g, 8.97 mmol), EDCI. HCl(2.33 g, 12.2 mmol) followed by HOBt (1.66 g, 12.24 mmol) and DIPEA (4.5mL, 24.4 mmol) at 0° C. The reaction mixture was warmed to RT andstirred for 16 h. After consumption of the starting material (by TLC),the reaction mixture was diluted with CH₂Cl₂. The separated organiclayer was washed with aqueous NaHCO₃ solution followed by aqueous NH₄Cl.The organic layer was dried over anhydrous Na₂SO₄, filtered andconcentrated under reduced pressure to get crude product, which waspurified by silica gel column chromatography eluting with 70%EtOAc/hexane to afford compound C-1 (2.3 g, 57.5%).

¹H-NMR: (400 MHz, DMSO-d₆): δ8.91 (d, 1H), 8.68 (d, 2H), 7.41 (br s,1H), 5.74 (br t, 1H), 5.07-4.89 (m, 1H), 4.15-4.10 (m, 1H), 3.97-3.92(m, 1H), 3.45-3.41 (m, 1H), 1.79-1.74 (m, 2H), 1.43-1.39 (m, 4H),1.29-1.21 (m, 6H), 1.12 (d, 5H), 0.71 (s, 9H), 0.12 (t, 1H), 0.09 (s,2H), 0.08 (s, 1H), 0.04 (s, 2H).

LCMS (m/z): 495.5 [M⁺+1].

Synthesis of tert-butyl2-(2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl)propyl)-1-oxo-2,5-diazaspiro[3.4]octane-5-carboxylate(C-2)

To a stirring solution of compound C-1 (2.3 g, 4.65 mmol) in THF (23 mL)was added TPP (1.34 g, 5.12 mmol) followed by DTAD (1.6 g, 6.98 mmol) at0° C. The reaction mixture was warmed to RT and stirred for 16 h. Afterconsumption of the starting material (by TLC), the reaction mixture wasconcentrated under reduced pressure to get crude product, which waspurified by silica gel column chromatography eluting with 25%EtOAc/hexane to afford compound C-2 (1.2 g, 54.2%)

¹H-NMR: (400 MHz, DMSO-d₆): δ8.82 (d, 2H), 7.49 (t, 1H), 4.72 (d, 1H),4.31 (q, 1H), 3.62 (br s, 2H), 3.25-3.19 (m, 1H), 2.24-2.05 (m, 2H),1.85-1.81 (m, 2H), 1.42 (br s, 1H), 1.25 (t, 3H), 0.92 (s, 8H), 0.75 (s,9H), 0.02 (s, 3H).

LCMS (m/z): 477.4 [M⁺+1].

Synthesis of tert-butyl 2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-1-oxo-2.5diazaspiro [3.4] octane-5-carboxylate (C-3)

To a stirring solution of compound C-2 (1.0 g, 2.10 mmol) in THF (20 mL)was added TBAF (1M in THF) (6.3 mL, 6.30 mmol) at 0° C. under N₂atmosphere and stirred for 1 h. After consumption of the startingmaterial (by TLC), the reaction mixture was quenched with ice water andextracted with EtOAc. The separated organic layer was dried overanhydrous Na₂SO₄, filtered and concentrated under reduced pressure toobtain crude product, which was purified by silica gel columnchromatography eluting with 5% MeOH/CH₂Cl₂ to afford compound C-3 (0.35g, 46%).

¹H-NMR: (400 MHz, DMSO-d₆): δ8.81 (d, 2H), 7.49 (t, 1H), 4.81 (d, 1H),4.65 (d, 1H), 4.25-4.20 (m, 1H), 3.64-3.51 (m, 2H), 3.34 (s, 1H),3.25-3.20 (m, 1H), 2.25-2.20 (m, 2H), 1.87-1.82 (m, 2H), 1.19 (d, 3H),0.97 (s, 9H).

LCMS (m/z): 363.3 [M⁺+1].

Synthesis of tert-butyl 2-(2-hydroxy-1-(pyrimidin-2-yl) propyl)-1-oxo-2,5-diazaspiro [3.4]octane-5-carboxylate (C-4)

To a stirring solution of compound C-3 (200 mg, 0.42 mmol) in THF (5 mL)was added TBAF (1M in THF) (0.84 mL, 0.84 mmol) at 0° C. under N₂atmosphere and stirred for 2 h. After consumption of the startingmaterial (by TLC), the reaction mixture was concentrated under reducedpressure to obtain crude product, which was purified by silica gelcolumn chromatography eluting with 2% MeOH/CH₂Cl₂ to afford C-4 (125 mg,82.2%).

¹H-NMR: (400 MHz, CD₃OD): 8.78 (t, J=3.2 Hz, 2H), 7.42-7.38 (m, 1H),4.92-4.87 (m, 1H), 4.79-4.56 (m, 1H), 4.43-4.38 (m, 1H), 4.33-3.97 (m,1H), 3.88-3.80 (m, 1H), 3.48-3.44 (m, 1H), 3.40-3.30 (m, 1H), 2.31-2.12(m, 2H), 1.96-1.84 (m, 2H), 1.42 (s, 9H), 1.32-1.28 (m, 3H);

LCMS (m/z): 363.3 [M⁺+1].

HPLC: 94.5%.

Synthesis of2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-2,5-diazaspiro[3.4]octan-1-one(C-5)

To a stirring solution of compound C-4 (0.3 g, 0.82 mmol) in CH₂Cl₂ (6mL) was added molecular sieves (0.3 g) followed by BF₃-etherate (0.31mL, 2.48 mmol) at 0° C. and stirred at RT for 1 h. After consumption ofthe starting material (by TLC), the reaction mixture was filtered andobtained residue was dissolved in MeOH and washed with CH₂Cl₂. Thevolatiles were evaporated under reduced pressure to obtain crudeproduct, which was purified by silica gel column chromatography elutingwith 8% MeOH/CH₂Cl₂ to afford C-5 (0.12 g, 55%).

¹H-NMR: (400 MHz, DMSO-d₆): δ8.81 (d, 2H), 7.49 (t, 1H), 4.81 (d, 1H),4.65 (d, 1H), 4.25-4.20 (m, 1H), 3.64-3.51 (m, 2H), 3.12-3.01 (m, 2H),2.15-2.10 (m, 2H), 1.87-1.82 (m, 2H), 1.19 (d, 3H).

LCMS (m/z): 263.1 [M⁺+1].

Synthesis of 2-(2-hydroxy-1-(pyrimidin-2-yl) propyl)-5-isobutyryl-2,5-diazaspiro [3.4]octan-1-one (C-6)

To a stirring solution of compound C-5 (500 mg, 1.9 mmol) in DCM (10 mL)was added TEA (0.0.79 mL, 5.72 mmol) followed by isobutyryl chloride(241 mg, 2.28 mmol) at 0° C. under N₂ atmosphere and stirred for 2 h atRT. After consumption of the starting material (by TLC), the reactionmixture was diluted with water (10 mL) and extracted with CH₂Cl₂ (2×15mL). The combined organic layer was washed with citric acid solution(1×20 mL). The separated organic layer was dried over anhydrous Na₂SO₄,filtered and concentrated under reduced pressure. Obtained crudematerial was purified by silica gel column chromatography eluting 2%MeOH/DCM to afford C-6 (85 mg, 13.4%) as white solid.

¹H-NMR: (400 MHz, CD₃OD): δ 4.91-4.75 (m, 1H), 4.46-4.40 (m, 1H),3.99-3.85 (m, 1H), 3.70-3.64 (m, 1H), 3.60-3.45 (m, 3H), 2.80-2.69 (m,1H), 2.29-2.22 (m, 2H), 2.07-1.96 (m, 2H), 1.45-1.33 (m, 3H), 1.16-1.10(m, 3H), 1.06-1.01 (m, 3H).

Synthesis of tert-butyl 2-(((1R, 2S)-2-((tert-butyldimethylsilyl)oxy)-1-(1, 3, 4-oxadiazol-2-yl) propyl) carbamoyl)-2-(hydroxymethyl)pyrrolidine-1-carboxylate (C-7)

To a stirring solution of compound I (2.1 g, 8.57 mmol) in CH₂Cl₂ (20mL) were added DIPEA (3.81 mL, 21.4 mmol), intermediate F (2.64 g, 10.28mmol), HATU (3.90 g, 10.28 mmol) at 0° C. and stirred to RT for 16 h.After consumption of the starting material (by TLC), the reactionmixture was diluted with water (30 mL). The organic layer was washedwith citric acid (1×50 mL) followed by brine solution (1×50 mL). Theorganic layer was dried over anhydrous Na₂SO₄, filtered and concentratedunder reduced pressure. Obtained crude material was purified by silicagel column chromatography eluting with 50% EtOAc/n-hexane to affordcompound C-7 (3.5 g, 85.15%) as yellow thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.24 (s, 1H), 8.17 (d, J=8.4 Hz, 1H),5.82-5.54 (m, 1H), 4.36-4.28 (m, 1H), 4.12-4.00 (m, 1H), 3.93-3.66 (m,1H), 3.43-3.39 (m, 1H), 2.69-2.66 (m, 2H), 2.02-1.94 (m, 2H), 1.78-1.68(m, 2H), 1.40 (s, 9H), 1.23-1.15 (m, 3H), 0.75 (s, 9H), −0.08 (s, 6H);

Mass (ESI): m/z 484.7 [M⁺+1]

Synthesis of tert-butyl 2-(((1R, 2S)-2-((tert-butyldimethylsilyl)oxy)-1-(1, 3, 4-oxadiazol-2-yl) propyl) carbamoyl)-2-(hydroxymethyl)pyrrolidine-1-carboxylate (C-8)

To a stirring solution of triphenylphosphine (4.05 g, 15.4 mmol) in THF(30 mL) was added DIAD (3.03 g, 15.4 mmol) at RT and stirred for 15 min.After added compound C-7 (3 g, 6.19 mmol) in (20 mL) THF slowly andreaction mixture was stirred at RT for 3 h. After consumption of thestarting material (by TLC), the reaction was concentrated under reducedpressure. The crude material was triturated with 30% Ether/n-hexane(2×50 mL). The filtered organic solvent was concentrated and cruderesidue was purified by column chromatography by eluting 30%EtOAc/n-hexane to afford compound C-8 (2 g, 69.4%) as thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.28 (s, 1H), 5.30-5.21 (m, 1H), 4.84-4.81(m, 1H), 4.49-4.29 (m, 1H), 4.05-3.91 (m, 1H), 3.65-3.61 (m, 1H),3.39-3.24 (m, 1H), 2.23-2.12 (m, 2H), 1.81-1.76 (m, 2H), 1.41 (s, 9H),1.20-1.14 (m, 3H), 0.75 (s, 9H), −0.22 (s, 6H);

Mass (ESI): m/z 467.6 [M⁺+1]

Synthesis of tert-butyl 2-((1R, 2S)-2-hydroxy-1-(1, 3, 4-oxadiazol-2-yl)propyl)-1-oxo-2, 5-diazaspiro [3.4] octane-5-carboxylate (C-9)

To a stirring solution of compound C-8 (2 g, 4.29 mmol) in THF (20 mL)was added TBAF (4.3 mL, 6.43 mmol) slowly at 0° C. and stirred at RT for30 min. After completion of reaction (by TLC), the reaction mixture wasevaporated under reduced pressure. The crude residue was purified bycolumn chromatography by eluting 70% EtOAc/n-hexane to afford C-9 (400mg, 26.5%) as white solid.

¹H-NMR: (400 MHz, DMSO-d₆): 9.29 (s, 1H), 5.30-5.21 (m, 1H), 5.08-4.95(m, 1H), 4.08-4.03 (m, 1H), 3.91-3.71 (m, 1H), 3.75-3.71 (m, 1H),3.65-3.52 (m, 1H), 3.36-3.22 (m, 1H), 2.22-2.09 (m, 2H), 1.83-1.77 (m,2H), 1.41 (s, 9H), 1.10-1.04 (m, 3H);

LCMS: 353.4;

HPLC: 99.82%.

Synthesis of 2-((1R, 2S)-2-hydroxy-1-(1, 3, 4-oxadiazol-2-yl)propyl)-2,5-diazaspiro [3.4]octan-1-one (C-10)

To a stirring solution of compound C-9 (500 mg, 1.07 mmol) in DCM (5 mL)was added BF₃(OEt)₂ (0.26 mL, 2.14 mmol) slowly at 0° C. and stirred atRT for 30 min. After completion of reaction (by TLC), the reactionmixture was evaporated under reduced pressure. The crude residue wastriturated with di ethylether/n-pentane (5 mL/5 mL) to obtain solid,C-10, which was taken into the next step without further purification.

Synthesis of 2-((1R, 2S)-2-hydroxy-1-(1, 3, 4-oxadiazol-2-yl)propyl)-5-isobutyryl-2, 5-diazaspiro [3.4] octan-1-one (C-11)

To a stirring solution of compound C-10 (700 mg, 2.77 mmol) in DCM (10mL) was added TEA (0.94 mL, 6.92 mmol), SM-3 (440 mg, 4.15 mmol) at 0°C. and stirred to RT for 1 h. After completion of reaction (by TLC),diluted with water (20 mL). The organic layer was washed with brinesolution (1×50 mL). The organic layer was dried over anhydrous Na₂SO₄,filtered and concentrated under reduced pressure. The crude residue waspurified by column chromatography followed by preparative HPLCpurification to afford C-11 (150 mg, 16.8%) as white solid.

¹H-NMR: (400 MHz, DMSO-d₆): 9.26 (s, 1H), 5.24-5.04 (m, 1H), 5.03-4.93(m, 1H), 4.27-3.91 (m, 1H), 3.90-3.64 (m, 1H), 3.49-3.45 (m, 3H),2.73-2.61 (m, 1H), 2.17-2.14 (m, 2H), 2.10-2.06 (m, 2H), 1.25-1.07 (m,3H), 0.95-0.93 (m, 6H).

LCMS m/z: 323.3.

HPLC: 97.89%.

Synthesis of 1-tert-butyl 2-methyl2-(hydroxymethyl)pyrrolidine-1,2-dicarboxylate (C-12)

To a stirring solution of compound G (74 g, 0.21 mol) in methanol (740mL) was added 10% Pd/C (50% wet, 14.8 g) under N₂ atmosphere and stirredfor 6 h under H₂ atmosphere (balloon pressure). The reaction mixture wasfiltered through celite pad and concentrated under reduced pressure toafford compound C-12 (45 g, 82%) as crude.

Synthesis of 1-tert-butyl 2-methyl 2-formylpyrrolidine-1,2-dicarboxylate(C-13)

To a stirring solution of compound C-12 (10 g, 38.5 mmol) in CH₂Cl₂ (100mL) was added Dess-Martin periodinane (19.6 g, 46.27 mmol) at 0° C.under N₂ atmosphere and stirred fro 3 h. After consumption of thestarting material (by TLC), the reaction was quenched with aqueousNaHCO₃ solution and extracted with CH₂Cl₂ (2×100 mL). The organic layerwas dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crudewas purified by column chromatography eluting with 10% EtOAc/Hexane toafford compound C-13 (7 g, 70.5%).

Synthesis of 1-tert-butyl 2-methyl2-((((1S,2R)-2-hydroxy-1-(1,2,4-oxadiazol-5-yl) propyl) amino)methyl)pyrrolidine-1,2-dicarboxylate (C-14)

To a stirring solution of compound C-13 (3 g, 11.6 mmol) in MeOH (30 mL)was added sodium acetate (1.91 g, 23.3 mmol) followed by intermediate S(3.6 g, 13.9 mmol). The reaction mixture was heated to reflux for 1 h.The reaction mixture was slowly cooled to RT-0° C., to this sodiumcyanoborohydride (1.465 g, 23.3 mmol) and stirring was continued foranother 6 h at RT. After consumption of the starting material (by TCL),methanol from the reaction was evaporated under reduced pressure and theresidue was diluted with water and extracted with EtOAc (2×50 mL). Theseparated organic layer was dried over anhydrous Na₂SO₄ and concentratedunder reduced pressure to give crude; which was purified by silica gelcolumn chromatography eluting with 40% EtOAc/Hexane to afford compoundC-14 (2.5 g, 56%).

LCMS m/z: 385 [M⁺+1].

Synthesis of tert-butyl 2-((1S,2R)-2-hydroxy-1-(1,2,4-oxadiazol-5-yl)propyl)-1-oxo-2,5-diazaspiro [3.4] octane-5-carboxylate (C-15)

To a stirring solution of compound C-14 (1.5 g, 3.90 mmol) in THF (30mL) was cooled to 0° C. and added t-BuMgCl (1M in THF, 15.6 mL, 15.6mmol) and stirred for 15 min. After consumption of the starting material(by TLC), the reaction mixture was quenched with aqueous NH₄Cl solutionand diluted with water. Aqueous layer was extracted with EtOAc (2×25mL). The separated organic layer was dried over anhydrous Na₂SO₄ andconcentrated under reduced pressure. Obtained crude material waspurified by silica gel column chromatography eluting with 150%EtOAc/CH₂Cl₂ to afford C-15 (0.15 g, 11%).

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.02 (s, 1H), 5.15 (s, 1H), 4.37-4.32 (m,1H), 3.95 (d, 1H), 3.66-3.60 (m, 1H), 3.36-3.30 (m, 1H), 2.29-2.09 (m,2H), 1.87-1.82 (m, 2H), 1.55 (s, 9H), 1.27 (d, 3H).

LCMS (ESI) m/z: 351 [M⁺−1].

HPLC Purity: 96%.

Synthesis of2-((1S,2R)-2-hydroxy-1-(1,2,4-oxadiazol-5-yl)propyl)-2,5-diazaspiro[3.4]octan-1-one (C-16)

To a stirring solution of C-15 (0.4 g, 1.13 mmol) in CH₂Cl₂ (4 mL) wasadded TFA (0.43 mL) at 0° C. and stirred at RT for 30 min. The reactionmixture was concentrated under vacuum. Obtained crude material waspurified by prep-HPLC to afford C-16 (65 mg) as TFA salt.

¹H-NMR: (400 MHz, DMSO-d6): δ 9.89 (br s, 1H), 9.08 (s, 1H), 5.46 (d,1H), 5.31 (s, 1H), 4.37-4.35 (m, 1H), 3.99 (d, 1H), 3.81 (d, 1H),3.42-3.35 (m, 2H), 2.35-2.18 (m, 2H), 2.10-2.03 (m, 2H), 1.24 (d, 3H).

LCMS (ESI) m/z: 253.4 [M⁺+1].

HPLC Purity: 95%.

Synthesis of tert-butyl 2-((2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl) propyl)carbamoyl)-2-(hydroxymethyl)-5-methylpyrrolidine-1-carboxylate (C-17)

To a stirring solution of compound Z (500 mg, 1.93 mmol) in DCM (10 mL)were added N,N-diisopropylethylamine (1.0 mL, 5.79 mmol), intermediate F(566 mg, 2.12 mmol), followed by EDCI (737 mg, 3.86 mmol). HOBT (521 mg,3.86 mmol) at 0° C. and stirred at RT for 16 h. After consumption of thestarting material (by TLC), the reaction mixture was diluted with water(20 mL). The separated organic layer was washed with saturated brinesolution (1×30 mL). The separated organic layer was dried over anhydrousNa₂SO₄ and concentrated under reduced pressure to afford crude compoundwhich was purified by column chromatography to obtained compound C-17(300 mg, 30.6%) as pale yellow liquid.

¹H-NMR: (500 MHz, CDCl₃): δ 8.73-8.68 (m, 3H), 5.30 (s, 2H), 4.40-4.15(m, 1H), 3.76-3.61 (m, 1H), 3.18-3.13 (m, 1H), 2.18-2.13 (m, 4H), 1.46(s, 9H), 1.32 (d, J=5.5 Hz, 3H), 1.24 (d, J=6.5 Hz, 3H), 1.08 (d, J=6.5Hz, 1H), 0.79 (s, 9H), 0.74 (t, J=10.5 Hz, 1H), 0.12 (s, 6H)

Synthesis of tert-butyl 2-(2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl) propyl)-6-methyl-1-oxo-2, 5-diazaspiro[3.4]octane-5-carboxylate (C-18)

To a stirring solution of triphenylphosphine (380 mg, 1.47 mmol) in dryTHF (10 mL) was added DTAD (339 mg, 1.47 mmol) at RT and stirred for 10min. After added compound C-17 (300 mg, 0.59 mmol) and the reactionmixture was stirred at RT for 16 h. After consumption of the startingmaterial (by TLC), the reaction mixture was concentrated under reducedpressure. The crude material was purified by silica gel columnchromatography eluting 30% EtOAc/hexane to afford compound C-18 (80 mg,27.5%) as yellow liquid.

¹H-NMR: (400 MHz. CDCl₃): 57.73-7.64 (m, 3H), 5.26 (s, 2H), 4.30 (dd,J=4.4 Hz, 4.4 Hz, 1H), 4.17-4.12 (m, 1H), 2.76 (t, J=6.8 Hz, 1H),2.32-2.28 (m, 2H), 2.05-2.00 (m, 2H), 1.49 (s, 9H), 1.45 (d, J=5.2 Hz,3H), 1.42 (d, J=4.8 Hz, 3H), 0.89 (s, 9H), 0.15 (s, 6H).

Synthesis of tert-butyl 2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-6-methyl-1-oxo-2, 5-diazaspiro [3.4] octane-5-carboxylate (C-19)

To a stirring solution of compound C-18 (700 mg, 1.42 mmol) in dry THF(10 mL) was added TBAF (744 mg, 2.85 mmol) at 0° C. The reaction mixturewas stirred at RT for 3 h. After consumption of the starting material(by TLC), the reaction mixture was concentrated under reduced pressure.The crude material was purified by silica gel column chromatographyeluting 80% EtOAc/hexane to afford C-19 (0.16 g, 30%) as yellow thicksyrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 8.77 (d, J=5.0 Hz, 2H), 7.42 (t, J=4.5 Hz,1H), 4.76-4.63 (m, 2H), 4.14-4.11 (m, 1H), 3.85-3.83 (m, 1H), 3.54-3.34(m, 1H), 2.25-2.21 (m, 2H), 2.08-1.93 (m, 1H), 1.53-1.47 (m, 1H), 1.39(s, 9H), 1.15 (d, J=6.0 Hz, 3H), 1.03-0.80 (m, 4H).

LCMS (ESI): 377.4 [M⁺+1].

HPLC: 94.6% (both isomers).

Synthesis of 2-(2-hydroxy-1-(pyrimidin-2-yl) propyl)-6-methyl-2,5-diazaspiro [3.4] octan-1-one (C-20)

To a stirring solution of compound C-19 (260 mg, 0.69 mmol) in DCM (10mL) was added 4A molecular sieves (70 mg), BF₃.OEt₂ (0.08 mL) at 0° C.under N₂ atmosphere. The reaction mixture was stirred at RT for 30 min.After consumption of the starting material (by TLC), the reactionmixture was concentrated under reduced pressure to afford crude, whichwas purified by preparative HPLC method to afford C-20 (65 mg, 34%) aswhite solid.

¹H-NMR: (400 MHz, D₂O): δ 8.88-8.86 (m, 2H), 7.63-7.57 (m, 1H),4.92-4.89 (m, 1H), 4.61-4.56 (m, 1H), 4.03-3.91 (m, 3H), 2.61-2.41 (m,3H), 1.92-1.84 (m, 1H), 1.46 (d, J=6.8 Hz, 3H), 1.37 (d, J=4.8 Hz, 3H);

LCMS (ESI): 295.3 [M⁺+H₂O].

HPLC: 91.2% (both enantiomers).

Synthesis of tert-butyl 2-((2-((tert-butyldimethylsilyl)oxy)-1-(1,3,4-oxadiazol-2-yl)propyl)carbamoyl)-2-(hydroxymethyl)-5-methylpyrrolidine-1-carboxylate (C-21)

To a stirring solution of compound Z (4 g, 15.4 mmol) in DCM (30 mL)were added N,N-diisopropylethylamine (8 mL, 46.33 mmol), intermediate N(4.74 g, 18.48 mmol) followed by HATU (7 g, 18.48 mmol) at 0° C. andstirred at RT for 12 h. After consumption of the starting material (byTLC), the reaction mixture was diluted with water (20 mL). The separatedorganic layer was washed with 10% citric acid solution (1×50 mL)followed by saturated brine solution (1×50 mL). The separated organiclayer was dried over anhydrous Na₂SO₄ and concentrated under reducedpressure to afford crude compound which was purified by columnchromatography eluting 30% EtOAc/n-hexane to obtained compound C-21 (5.5g, 71.8%) as pale yellow liquid.

¹H-NMR: (500 MHz, DMSO-d₆): 59.25 (s, 1H), 5.50-5.41 (m, 1H), 5.19-5.06(m, 2H), 4.38-4.36 (m, 1H), 4.05-3.90 (m, 3H), 2.69-1.98 (m, 2H), 1.39(s, 9H), 1.37-1.26 (m, 2H), 1.23-1.16 (m, 6H), 0.76 (s, 9H), −0.01 (s,6H).

Synthesis of tert-butyl2-(2-((tert-butyldimethylsilyl)oxy)-1-(1,3,4-oxadiazol-2-yl)propyl)-6-methyl-1-oxo-2,5-diazaspiro[3.4]octane-5-carboxylate(C-22)

To a stirring solution of triphenylphosphine (5.5 g, 11.04 mmol) in THF(55 mL) was added DTAD (5.07 g, 22.05 mmol) at RT and stirred for 10min. After added compound C-21 (5.5 g, 1.04 mmol) and the reactionmixture was stirred at RT for 16 h. After consumption of the startingmaterial (by TLC), the reaction mixture was concentrated under reducedpressure. The crude material was purified by silica gel columnchromatography eluting 30% EtOAc/hexane to afford compound C-22 (4 g,75.4%) as yellow liquid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.36 (s, 1H), 5.30-4.48 (m, 1H), 4.47-4.31(m, 1H), 4.03-3.88 (m, 2H), 3.62-3.58 (m, 1H), 2.38-1.98 (m, 3H),1.57-1.53 (m, 1H), 1.41 (s, 9H), 1.38-1.15 (m, 6H), 0.69 (s, 9H), −0.01(s, 6H).

Synthesis tert-butyl2-(2-hydroxy-1-(1,3,4-oxadiazol-2-yl)propyl)-6-methyl-1-oxo-2,5-diazaspiro[3.4] octane-5-carboxylate (C-23)

To a stirring solution of compound C-22 (3.5 g, 7.29 mmol) in THF (35mL) was added TBAF (1M THF) (14.5 mL, 14.58 mmol) at 0° C. The reactionmixture was stirred at RT for 1 h. After consumption of the startingmaterial (by TLC), the reaction mixture was concentrated under reducedpressure. The crude material was purified by silica gel columnchromatography eluting 60% EtOAc/hexane to afford C-23 (800 mg, 30.7%)as an off-white solid.

¹H-NMR: (400 MHz, DMSO-d₆): δ9.30 (s, 1H), 5.30-5.21 (m, 1H), 5.13-5.02(m, 1H), 4.13-4.03 (m, 1H), 3.90-3.83 (m, 1H), 3.74-3.68 (m, 1H),3.62-3.49 (m, 1H), 2.29 (m, 3H), 1.56-1.52 (m, 1H), 1.40 (s, 9H),1.25-1.14 (m, 6H).

LCMS (ESI) (m/z): 377.4 [M⁺+1].

HPLC: 95.38%.

Synthesis of tert-butyl 2-(((1R, 2S)-2-((tert-butyldimethylsilyl)oxy)-1-(1, 3, 4-oxadiazol-2-yl) propyl) carbamoyl)-2-(1-hydroxyethyl)pyrrolidine-1-carboxylate (C-24)

To a stirring solution of compound BB (3 g, 11.5 mmol) in CH₂Cl₂ (30 mL)were added DIPEA (6 mL, 34.5 mmol), intermediate N (2.95 g, 11.5 mmol).HATU (5.24 g, 13.8 mmol) at 0° C. and stirred to RT for 12 h. Afterconsumption of the starting material (by TLC), the reaction mixture wasdiluted with water (30 mL) and extracted with CH₂Cl₂ (50 mL). Thecombined organic layer was washed with citric acid (1×50 mL) followed bybrine solution (1×50 mL). The organic layer was dried over anhydrousNa₂SO₄, filtered and concentrated under reduced pressure. Obtained crudematerial was purified by silica gel column chromatography eluting with35% EtOAc/n-hexane to afford compound C-24 (3.6 g, 62.5%) as yellowthick syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.26 (s, 1H), 8.74 (d, J=8.5 Hz, 2H),5.30-5.12 (m, 1H), 4.61-4.58 (m, 1H), 4.38-4.35 (m, 2H), 3.57-3.54 (m,1H), 2.68-1.98 (m, 2H), 1.97-1.69 (m, 2H), 1.40 (s, 9H), 1.19-1.00 (m,6H), 0.85 (s, 6H), −0.25 (s, 9H).

Mass (ESI): m/z 497.6 [M⁻−1].

Synthesis of tert-butyl 2-((1R, 2S)-2-((tert-butyldimethylsilyl)oxy)-1-(1, 3, 4-oxadiazol-2-yl) propyl)-1-methyl-3-oxo-2, 5-diazaspiro[3.4]octane-5-carboxylate (C-25)

To a stirring solution of triphenylphosphine (3.6 g, 14.05 mmol) in THF(20 mL) was added DIAD (2.8 g, 14.05 mmol) at RT and stirred for 30 min.After added compound C-24 (3.5 g, 7.02 mmol) in (15 mL) THF slowly andreaction mixture was stirred at RT for 6 h. After consumption of thestarting material (by TLC), the reaction was concentrated under reducedpressure. The crude material was purified by column chromatography byeluting 30% EtOAc/n-hexane to afford compound C-25 (3 g, 89.2%) as thicksyrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.33 (s, 1H), 5.33-5.29 (m, 2H), 4.79-4.75(m, 1H), 4.50-4.45 (m, 1H), 4.06-3.96 (m, 1H), 2.16-2.08 (m, 1H),2.01-1.89 (m, 1H), 1.81-1.76 (m, 2H), 1.40 (s, 9H), 1.24-1.15 (s, 6H),0.80 (s, 9H). −0.08 (s, 6H).

Mass (ESI): m/z 319.3 [M⁺+1].

Synthesis of tert-butyl 2-((1R, 2S)-2-hydroxy-1-(1, 3, 4-oxadiazol-2-yl)propyl)-1-methyl-3-oxo-2, 5-diazaspiro [3.4]octane-5-carboxylate (C-26)

To a stirring solution of compound C-25 (3 g, 6.25 mmol) in THF (25 mL)was added TBAF (12.5 mL, 12.5 mmol) at 0° C. and stirred for 30 min.After consumption of the starting material (by TLC), the reactionmixture was concentrated under reduced pressure. Obtained crude materialwas purified by silica gel column chromatography eluting with 60%EtOAc/n-hexane to afford C-26 (0.8 g, 35%) as white solid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.29 (s, 1H), 5.28-5.15 (m, 1H), 5.08-4.89(m, 1H), 4.43-4.35 (m, 1H), 3.97-3.79 (m, 1H), 3.65-3.40 (m, 1H),3.31-3.25 (m, 1H), 2.15-2.00 (m, 2H), 1.93-1.86 (m, 1H), 1.80-1.68 (m,1H), 1.40 (s, 9H), 1.18-1.04 (m, 6H);

Mass (ESI): nm/z 367.4 [M⁺+1].

HPLC: 95.85% (both isomers).

Synthesis of tert-butyl 2-((2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl) propyl) carbamoyl)-2-(1-hydroxyethyl)pyrrolidine-1-carboxylate (C-27)

To a stirring solution of compound BB (200 mg, 0.77 mmol) in CH₂Cl₂ (20mL) were added DIPEA (298 mg, 2.31 mmol). EDCI (221 mg, 1.15 mmol). HOBt(177 mg, 1.15 mmol) followed by intermediate F (206 mg, 0.77 mmol) at 0°C. and stirred for 16 h at RT. After consumption of the startingmaterial (by TLC), the reaction mixture was concentrated under reducedpressure. Obtained crude material was purified by silica gel columnchromatography eluting with 4% MeOH/DCM to afford compound C-27 (210 mg,53.6%) as pale yellow thick syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 8.77 (d, J=4.5 Hz, 2H), 7.40 (t, J=5.0 Hz,1H), 4.95-4.92 (m, 2H), 4.21-4.05 (m, 2H), 3.34-3.25 (m, 2H), 3.20-3.12(m, 1H), 1.81-1.63 (m, 4H), 1.48 (s, 9H), 1.12-1.06 (m, 3H), 0.99-0.85(m, 3H), 0.70 (s, 9H), −0.06 (s, 6H);

LCMS (m/z): 509.4 [M⁺+1]

Synthesis of tert-butyl 2-(2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl) propyl)-1-methyl-3-oxo-2, 5-diazaspiro[3.4]octane-5-carboxylate (C-28)

To a stirring solution of triphenylphosphine (206 mg, 0.78 mmol) in THF(6 mL) was added DIAD (179 mg, 0.78 mmol) at RT and stirred for 30 min.To this added compound C-27 (200 mg, 0.39 mmol) in (5 mL) THF slowly andreaction mixture was stirred at RT for 2 h. After consumption of thestarting material (by TLC), the reaction was concentrated under reducedpressure. The crude material was triturated with diethylether/n-pentane(10 mL/10 mL). The filtered solvent was concentrated and purified bysilica gel column chromatography eluting 30% EtOAc/n-hexane to affordcompound C-28 (80 mg, 41.8%) as pale yellow liquid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.81-8.77 (m, 2H), 7.72 (d, J=5.5 Hz, 1H),4.92-4.80 (m, 1H), 4.70-4.65 (m, 2H), 4.46-4.20 (m, 1H), 3.30-3.16 (m,1H), 1.86-1.65 (m, 2H), 1.64-1.43 (m, 2H), 1.40 (s, 9H), 1.20-1.01 (m,3H), 0.92-0.85 (m, 3H), 0.74 (s, 9H), −0.06 (s, 6H);

LCMS (m/z): 491.4 [M⁺+1].

Synthesis of tert-butyl 2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-1-methyl-3-oxo-2, 5-diazaspiro [3.4] octane-5-carboxylate (C-29)

To a stirring solution of compound C-28 (450 mg, 0.91 mmol) in THF (10mL) was added TBAF (1.8 mL, 1.83 mmol) in 5 mL THF at 0° C. and stirredto RT for 4 h. After consumption of the starting material (by TLC), thereaction was concentrated under reduced pressure. The crude material waspurified by silica gel column chromatography eluting 2% MeOH/DCMfollowed by preparative HPLC purification to afford C-29 (100 mg, 29%)as a semi solid.

¹H-NMR: (400 MHz, CD₃OD): δ 8.82-8.79 (m, 2H), 8.78-7.43 (m, 1H),4.94-4.89 (m, 1H), 4.63-4.44 (m, 1H), 3.99-3.73 (m, 1H), 3.56-3.36 (m,2H), 2.29-2.22 (m, 2H), 1.99-1.73 (m, 2H), 1.40 (s, 9H), 1.24-1.20 (m,3H), 1.13-1.05 (m, 3H).

LCMS (m/s): 377.5 [M⁺+1].

HPLC: 94.4% (both isomers).

Synthesis of tert-butyl 2-((2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl) propyl)carbamoyl)-2-(1-hydroxyethyl)-5-methylpyrrolidine-1-carboxylate (C-30)

To a stirring solution of compound DD (500 mg, 1.83 mmol) in DCM (10 mL)were added N,N-diisopropylethylamine (0.84 mL, 4.57 mmol), intermediateF (586 mg, 2.19 mmol), followed by EDCI (296 mg, 2.19 mmol), HOBT (338mg, 2.19 mmol) at 0° C. and stirred at RT for 16 h. After consumption ofthe starting material (by TLC), the reaction mixture was diluted withwater (20 mL). The separated organic layer was washed with brinesolution (1×20 mL). The separated organic layer was dried over anhydrousNa₂SO₄ and concentrated under reduced pressure to afford crude compoundwhich was purified by column chromatography by eluting 2% MeOH/DCM toobtained compound C-30 (800 mg, 83.6%) as pale yellow liquid.

¹H-NMR: (500 MHz, DMSO-d₆): δ8.78-8.75 (m, 2H), 7.42-7.39 (m, 1H),5.01-4.92 (m, 1H), 4.21-4.09 (m, 1H), 4.05-4.02 (m, 2H), 1.98 (s, 2H),1.44-1.31 (m, 7H), 1.29-1.11 (m, 10H), 1.02-0.96 (m, 4H), 0.70-0.63 (m,9H), −0.25 (s, 3H), −0.32 (s, 3H).

LCMS (ESI): 523.6 [M⁺+1].

Synthesis of tert-butyl 2-(2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl) propyl)-1, 6-dimethyl-3-oxo-2, 5-diazaspiro[3.4]octane-5-carboxylate (C-31)

To a stirring solution of triphenylphosphine (1.23 g, 4.87 mmol) in dryTHF (10 mL) was added DTAD (1.09 g, 4.77 mmol) as portion-wise andstirred for 15 min at RT. To this precipitated solution, added compoundC-30 (1.0 g, 1.91 mmol) in dry THF (10 mL) slowly at RT and stirred for16 h. After consumption of the starting material (by TLC), the reactionmixture was concentrated under reduced pressure. The crude material wastriturated with n-pentane (20 mL) and filtered solid (TPPO). Thefiltrate was concentrated under reduced pressure to obtained crudecompound was purified by silica gel column chromatography eluting 30%EtOAc/hexane to afford compound C-31 (400 mg, 41.4%) as yellow liquid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.82-8.79 (m, 2H), 7.44-7.42 (m, 1H),3.88-3.70 (m, 2H), 1.82-1.78 (m, 2H), 1.49-1.41 (m, 4H), 1.40 (s, 9H),1.21-1.08 (m, 9H), 0.57 (s, 6H), −0.05 (s, 9H);

LCMS (ESI): 505.5 [M⁺+1].

Synthesis of tert-butyl 2-(2-hydroxy-1-(pyrimidin-2-yl) propyl)-1,6-dimethyl-3-oxo-2, 5-diazaspiro [3.4] octane-5-carboxylate (C-32)

To a stirring solution of compound C-31 (220 mg, 0.43 mmol) in dry THF(3 mL) was added TBAF (227 mg) slowly at 0° C. and stirred at RT for 16h. After consumption of the starting material (by TLC), the reactionmixture was concentrated under reduced pressure to afford crude, whichwas purified by silica gel column chromatography eluting 80%EtOAc/hexane to afford compound C-32 (150 mg, 88.2%) as yellow liquid.

¹H-NMR: (500 MHz, DMSO-d₆): 58.81-8.79 (m, 2H), 7.43-7.41 (m, 1H),4.36-4.33 (m, 1H), 4.27-4.24 (m, 1H), 3.86-3.80 (m, 2H), 2.15-1.86 (m,4H), 1.53-1.50 (m, 1H), 1.43 (s, 9H), 1.19-1.03 (m, 9H).

Synthesis of 2-(2-hydroxy-1-(pyrimidin-2-yl) propyl)-3, 6-dimethyl-2,5-diazaspiro [3.4]octan-1-one (C-33)

To a stirring solution of compound C-32 (120 mg, 0.25 mmol) in DCM (10mL) was added molecular sieves (100 mg), BF₃(OEt)₂ (72 mg, 0.51 mmol) at0° C. under N₂ atmosphere. The reaction mixture was stirred at RT for 10min. After consumption of the starting material (by TLC), the obtainedprecipitate was triturated with n-pentane/diethylether (5 mL/5 mL) andthe filtered solid was dried on vacuum to afford C-33 (80 mg, 89.8%) aswhite solid.

¹H-NMR: (400 MHz. D₂O): δ 8.90 (t, J=4.8 Hz, 2H), 7.64-7.60 (m, 1H),4.72-4.62 (m, 2H), 4.24-4.15 (m, 1H), 4.08-4.00 (m, 1H), 2.60-2.32 (m,3H), 2.02-1.91 (m, 1H), 1.55-1.51 (m, 3H), 1.48-1.45 (m, 3H), 1.39-1.25(m, 3H).

LCMS (ESI): 581.3 [2M⁺+1].

Synthesis of tert-butyl 2-((2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl) propyl) carbamoyl)-2-(1-hydroxyethyl)pyrrolidine-1-carboxylate (C-34)

To a stirring solution of compound BB (2.5 g, 9.65 mmol) in CH₂Cl₂ (25mL) were added DIPEA (5 mL, 28.95 mmol), intermediate F (2.57 g, 9.65mmol) followed by HATU (4.4 g, 11.58 mmol) at 0° C. and stirred for 12 hat RT. After consumption of the starting material (by TLC), the reactionmixture was the reaction was diluted with water (20 mL). The separatedorganic layer was washed with citric acid solution (1×50 mL) followed bybrine solution (1×50 mL). The organic layer was dried over anhydrousNa₂SO₄, filtered and concentrated under reduced pressure. Obtained crudematerial was purified by silica gel column chromatography eluting 50%EtOAc/n-hexane to afford compound C-34 (3.1 g, 63.2%) as an off-whitesolid.

¹H-NMR: (500 MHz, DMSO-d₆): δ 8.76 (d, J=13.5 Hz, 2H), 8.42 (d, J=9.0Hz, 1H), 7.41 (d, J=9.0 Hz, 1H), 4.60-4.41 (m, 1H), 4.23-4.11 (m, 1H),3.58-3.40 (m, 3H), 2.37-1.62 (m, 4H), 1.39 (s, 9H), 1.28-1.19 (m, 3H),1.18-1.15 (m, 3H), 1.09 (s, 9H), −0.04 (s, 6H);

LCMS (ESI): m/z 509.7 [M⁺+1]

Synthesis of tert-butyl 2-(2-((tert-butyldimethylsilyl)oxy)-1-(pyrimidin-2-yl) propyl)-1-methyl-3-oxo-2, 5-diazaspiro[3.4]octane-5-carboxylate (C-35)

To a stirring solution of triphenylphosphine (1.6 g, 6.29 mmol) in THF(15 mL) was added DIAD (1.44 g, 6.29 mmol) at RT and stirred for 30 min.To this added compound C-34 (1.6 g, 3.14 mmol) in (5 mL) THF slowly andreaction mixture was stirred at RT for 3 h. After consumption of thestarting material (by TLC), the reaction was concentrated under reducedpressure. The crude material was triturated with diethyl ether/n-pentane(10 mL/10 mL). The filtered solvent was concentrated and purified bysilica gel column chromatography eluting 20% EtOAc/n-hexane to affordcompound C-35 (1 g, 65.3%) as pale yellow liquid.

¹H-NMR: (500 MHz, DMSO-d₆): δ 8.80 (d, J=14.5 Hz, 2H), 7.44 (d, J=9.0Hz, 1H), 4.72-4.46 (m, 2H), 3.36-3.23 (m, 3H), 2.06-1.61 (m, 4H), 1.40(s, 9H), 1.35-1.23 (m, 3H), 1.16-1.12 (m, 3H), 1.00 (s, 9H), −0.01 (s,6H);

LCMS (ESI): m/z 491.4 [M⁺+1]

Synthesis of 2-(2-hydroxy-1-(pyrimidin-2-yl) propyl)-3-methyl-2,5-diazaspiro [3.4] octan-1-one (C-36)

To a stirring solution of compound C-35 (1 g, 2.04 mmol) in DCM (5 mL)was added BF₃O(C₂H5)₂ (0.05 mL, 4.08 mmol) followed by molecular sieves(50 mg) at 0° C. and stirred to RT for 3 h. After consumption of thestarting material (by TLC), the reaction was diluted with n-pentane andfiltered the obtained solid. The filtered solid was purified by silicagel column chromatography eluting 5% MeOH/DCM to afford C-36 (180 mg,32%) as an off-white solid.

¹H-NMR: (400 MHz, D₂O): δ 8.89 (d, J=4.8 Hz, 2H), 7.61 (t, J=4.8 Hz,1H), 4.85-4.73 (m, 2H), 4.18-4.13 (m, 1H), 3.54-3.41 (m, 2H), 2.43-2.36(m, 1H), 2.31-2.12 (m, 3H), 1.43-1.37 (m, 6H)

LCMS (ESI): m/z 277.3 [M⁺+1];

HPLC: 98.85%.

Synthesis of 2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-5-isobutyryl-3-methyl-2, 5-diazaspiro [3.4] octan-1-one (C-37)

To a stirring solution of compound C-36 (600 mg, 2.17 mmol) in CH₂Cl₂(10 mL) was added TEA (1.05 mL, 7.59 mmol) at 0° C. After addedisobutyryl chloride (253 mg, 2.39 mmol) slowly and stirred for 1 h atRT. After consumption of the starting material (by TLC), the reactionmixture was diluted with water (10 mL). The separated organic layer waswashed with brine solution (1×50 mL). The organic layer was dried overanhydrous Na₂SO₄, filtered and concentrated under reduced pressure.Obtained crude material was purified by silica gel column chromatographyeluting 3% MeOH/DCM followed by preparative HPLC purification to affordC-37 (120 mg, 16%) as an off-white solid.

¹H-NMR: (500 MHz, DMSO-d₆): δ 8.81 (d, J=7.5 Hz, 2H), 7.43 (d, J=11.5Hz, 1H), 5.05-4.34 (m, 2H), 3.64-3.44 (m, 3H), 2.01-1.81 (m, 5H),1.35-1.13 (m, 6H), 1.10-1.04 (m, 6H);

LCMS (ESI): m/z 347.4 [M⁺+1]

HPLC: 91.1%

Preparation of Key Intermediates, Schemes I-8 to I-10:

Synthesis of ethyl 2-((benzyloxy)methyl)-5-methylpyrrolidine-2-carboxylate (EE)

To a stirring solution of X (8.5 g, 22.51 mmol) in DCM (50 mL) was addedTFA (8.6 mL, 112.58 mmol) at 0° C. The reaction mixture was stirred atRT for 4 h. After consumption of the starting material (by TLC), thereaction mixture was concentrated under reduced pressure to afford EE(14 g, crude) as pink liquid (TFA salt).

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.71 (br s, 1H), 7.45-7.30 (m, 5H), 4.62(s, 2H), 4.27-4.22 (m, 2H), 3.90-3.73 (m, 3H), 2.26-2.08 (m, 2H),2.01-1.95 (m, 1H), 1.59-1.51 (m, 1H), 1.36-1.31 (m, 3H), 1.22-1.18 (m,3H);

LCMS (ESI): m/z 278.36 [M⁺+1]

Synthesis of ethyl 2-(hydroxymethyl)-5-methylpyrrolidine-2-carboxylate(FF)

To a stirring solution of EE (14 g (crude), 35.80 mmol) in methanol (100mL) was added (50% wet) 10% Pd/C (5 g) under N₂ atmosphere. The reactionmixture was stirred under H₂ atmosphere (balloon pressure) at RT for 16h. After consumption of the starting material (by TLC), the reactionmixture was filtered through a pad of celite and the pad was washed withmethanol (100 mL). Obtained filtrate was concentrated under reducedpressure to afford FF (8 g, crude) as yellow syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.52 (s, 1H), 8.74 (s, 1H), 4.28-4.20 (m,2H), 3.94-3.69 (m, 3H), 2.22-2.16 (m, 2H), 2.13-1.92 (m, 1H), 1.60-1.53(m, 1H), 1.32-1.18 (m, 6H);

LCMS (ESI): m/z 188.24 [M⁺+1]

Synthesis of ethyl1-benzyl-2-(hydroxymethyl)-5-methylpyrrolidine-2-carboxylate (GG)

To a stirring solution of FF (8 g, 26.57 mmol) in CH₃CN (50 mL) wasadded K₂CO₃ (11.02 g, 79.73 mmol) followed by benzyl bromide (3.78 mL,31.89 mmol) at RT and stirred for 16 h. After consumption of thestarting material, filtered the reaction mass through celite bed andfilterate was washed with EtOAc (250 mL). The filterate was concentratedunder reduced pressure to afford crude. Obtained crude material waspurified by column chromatography eluting with 10% EtOAc/Hexane toafford GG (5 g, 68%) as brown syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 7.33-7.15 (m, 5H), 5.17 (t, J=5.5 Hz, 1H),4.75 (d, J=5.5 Hz, 1H), 4.50 (d, J=6.0 Hz, 1H), 4.14-3.93 (m, 2H), 3.93(s, 2H), 3.81-3.60 (m, 1H), 3.13-3.09 (m, 1H), 2.20-2.17 (m, 1H),1.99-1.88 (m, 2H), 1.40-1.36 (m, 3H), 1.14 (t, J=7.0 Hz, 3H)

LCMS (ESI): m/z 278.36 [M⁺+1]

Synthesis of 1-benzyl-2-(hydroxymethyl)-5-methylpyrrolidine-2-carboxylicAcid (HH)

To a stirring solution of GG (5 g, 18.05 mmol) in EtOH/H₂O (25 mL/25 mL)were added NaOH (1.44 g, 36.1 mg) at RT. The reaction mixture was heatedto 100° C. for 2 h. After consumption of the starting material (by TLC),the solvent from the reaction was evaporated under reduced pressure andextracted with di ethylether (2×75 mL). The aqueous layer was acidifiedby using 1N HCl and extracted with 20% MeOH/CH₂Cl₂ (2×100 mL). Theorganic layer was dried over anhydrous Na₂SO₄, filtered and concentratedunder reduced pressure to afford HH (3.5 g, 78%) as brown syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 11.17 (s, 1H), 7.49-7.27 (m, 5H), 4.55 (d,J=14.0 Hz, 1H), 4.06 (d, J=14.4 Hz, 1H), 3.83 (s, 2H), 3.51-3.41 (m,2H), 2.18-1.90 (m, 3H), 1.45-1.39 (m, 1H), 0.61-0.45 (m, 3H)

LCMS (ESI): m/z 250.13 [M⁺+1]

Synthesis of methyl pyrrolidine-2-carboxylate (II):

To a stirring solution of L-Proline (50 g, 434 mmol) in methanol wasadded thionyl chloride (37.5 ml, 521 mmol) at 0° C. and heated to 70° C.for 16 h. The reaction mixture was brought to RT and concentrated undervacuum to afford II as (70 g, 99%) as thick syrup (hydrochloride salt).

¹H-NMR: (500 MHz, DMSO-d₆): δ 4.15-4.13 (m, 1H), 3.65 (s, 3H), 3.35-3.30(m, 2H), 2.23-2.15 (m, 1H), 1.86-1.78 (m, 3H), 1.41 (s, 9H)

LCMS (ESI): m/z 129 [M⁺+1]

Synthesis of methylbenzylprolinate (JJ)

To a stirring solution of compound II (18 g, 108 mmol) in DCM (200 mL)was added TEA (45.35 mL, 326 mmol) followed by benzyl bromide (15.5 mL,130 mmol) at 0° C. and stirred at RT for 24 h. After completion of thereaction (by TLC) was diluted with water (75 mL) and EtOAc (0.500 mL).The organic layer was washed with water (2×100 mL), brine solution (2×50mL). The organic layer was dried over anhydrous Na₂SO₄ and concentratedunder vacuum. Obtained crude material was purified by columnchromatography eluting with 10% EtOAc/Hexane to afford JJ (5 g, 21%) asoily liquid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 7.32-7.22 (m, 5H), 3.86-3.80 (m, 1H), 3.58(s, 3H), 3.48-3.24 (m, 2H), 2.85-2.80 (m, 1H), 2.38-2.32 (m, 1H),2.10-1.98 (m, 1H), 1.85-1.69 (m, 3H)

LCMS (ESI): m/z 220.28 [M⁺+1]

Synthesis of methyl 1-benzyl-2-(1-hydroxyethyl)pyrrolidine-2-carboxylate (KK)

To a stirring solution of JJ (5 g, 22.80 mmol) in THF (50 mL) was addedLiHMDS (46 mL, 45.60 mmol) dropwise at −20° C. and stirred for 45 min.To this acetaldehyde (2.33 mL, 45.60 mmol) was added drop wise andstirred for 3 h at −20° C. After consumption of the starting material(by TLC), the reaction was quenched with aqueous NH₄Cl solution (100 mL)and extracted with EtOAc (2×150 mL). The separated organic layer waswashed with water (200 mL) and brine solution (200 mL). The separatedorganic layer was dried over Na₂SO₄ and concentrated to afford crudewhich was purified by column chromatography eluting with 10%EtOAc/Hexane to afford KK (3.5 g, 58%) as yellow syrup.

LCMS (ESI): m/z 264.3 [M⁺+1]

Synthesis of 1-benzyl-2-(1-hydroxyethyl) pyrrolidine-2-carboxylic Acid(LL)

To a stirring solution of KK (4.5 g, 17.11 mmol) in MeOH/THF/H₂O (10mL/10 mL/10 mL) were added NaOH (1.02 g, 25.66 mmol) at RT. The reactionmixture was heated to 95° C. for 1 h. After consumption of the startingmaterial (by TLC), the solvent was evaporated under reduced pressure.The aqueous layer was washed with EtOAc (100 mL). The separated aqueouslayer was acidified by using 1N HCl (pH˜3). The aqueous layer wasextracted with 20% MeOH/DCM (2×100 mL). The combined organic layer wasdried over Na₂SO₄ and concentrated under reduced pressure to afford LL(2.7 g, 63%) as an off-white solid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 7.31-7.13 (m, 5H), 3.87-3.78 (m, 2H),3.61-3.57 (m, 1H), 2.75-2.64 (m, 2H), 1.90-1.78 (m, 2H), 1.69-1.65 (m,1H), 1.53-1.43 (m, 1H), 1.06 (d, J=6.4 Hz, 3H);

LCMS (ESI): m/z 250.4 [M⁺−1]

Synthesis of methyl L-serinate (MM)

To a stirring solution of L-serine (12 g, 114 mmol) in CH₃OH (100 mL)was added thionyl chloride (10 mL, 137 mmol) at 0° C. and stirred at 80°C. for 16 h. After completion of starting material (by TLC), thereaction mixture was concentrated under reduced pressure to afford MM(16 g. crude, HCl salt) as white solid. This material was directly usedfor the next step without further purification.

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.61 (s, 2H), 4.50 (s, 1H), 4.20-4.16 (m,1H), 3.82 (d, J=3.6 Hz, 2H), 3.73 (s, 3H)

Synthesis of methyl ((benzyloxy)carbonyl)-L-serinate (NN)

To a stirring solution of MM (16 g, 103 mmol) in water/1, 4 dioxane (130mL/65 mL) were added added Na₂CO₃ (27.3 g, 257 mmol) at RT. After addedCbz-Cl (17.6 mL, 123 mmol) was added at 0° C. drop wise and stirred for16 h at RT. After completion of starting material (by TLC), diluted thereaction mass with EtOAc (300 ml). The separated organic layer was driedover anhydrous Na₂SO₄, filtered and concentrated under reduced pressureto afford crude material which was purified by column chromatography toafford NN (23 g, 88%) as thick syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 7.50 (d, J=8.0 Hz, 1H), 7.38-7.31 (m, 5H),5.04 (s, 2H), 4.95-4.92 (m, 1H), 4.17-4.13 (m, 1H), 3.66 (s, 3H),3.65-3.56 (m, 2H).

LCMS (ESI): m/z 254.2 [M⁺+1]

Synthesis of methylN-((benzyloxy)carbonyl)-O-(tert-butyldimethylsilyl)-L-serinate (OO)

To a stirring solution of NN (23 g, 91 mmol) in DCM (700 mL) were addedimidazole (12.37 g, 182 mmol). DMAP (2.22 g, 18.2 mmol) followed byTBDMS-Cl (20.4 g, 136 mmol) at 0° C. and stirred at RT for 16 h. Aftercompletion of starting material (by TLC), diluted the reaction mass withwater (0.200 ml). The separated organic layer was washed with brinesolution (2×200 mL). The organic layer was dried over anhydrous Na₂SO₄,filtered and concentrated under reduced pressure to afford crudematerial was purified by column chromatography eluting 20% EtOAc/hexaneto afford OO (32 g, 97%) as thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 7.56 (d, J=8.0 Hz, 1H), 7.38-7.30 (m, 5H),5.07 (s, 2H), 4.24-4.19 (m, 1H), 3.81 (d, J=5.2 Hz, 2H), 3.63 (s, 3H),0.84 (s, 9H), −0.04 (s, 6H)

LCMS (ESI): m/z 368.5 [M⁺+1]

Synthesis of benzyl(S)-(3-((tert-butyldimethylsilyl)oxy)-1-hydrazinyl-1-oxopropan-2-yl)carbamate(PP)

To a stirred solution of OO (20 g, 54.5 mmol) in methanol (120 mL) wasadded hydrazine hydrate (27 g, 545 mmol) at 0° C. and after stirred at80° C. for 2 h. After completion of starting material (by TLC), ethanolwas evaporated under reduced pressure. The crude residue was trituratedwith n-pentane (100 mL) to afford PP (17 g, 85%) as an off-white solid.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.17 (s, 1H), 9.10 (s, 1H), 7.35-7.14 (m,5H), 5.02 (s, 2H), 4.48 (s, 2H), 4.16-4.07 (m, 2H), 3.73-3.58 (m, 1H),0.81 (s, 9H), −0.04 (s, 6H).

Synthesis of benzyl ((1S, 2R)-2-((tert-butyldimethylsilyl) oxy)-1-(1, 3,4-oxadiazol-2-yl) propyl) carbamate (QQ)

A solution of PP (17 g, 46.3 mmol) in triethylorthoformate (68.5 g, 463mmol) was added p-TSA (8.8 g, 4.63 mmol) at RT and after stirred at 80°C. for 2 h. After completion of starting material (by TLC),triethylorthoformate was evaporated under reduced pressure. The cruderesidue was purified by column chromatography eluting 10% EtOAc/hexaneto afford QQ (5 g, 29%) as thick syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.22 (s, 1H), 8.05 (d, J=7.5 Hz, 1H),7.36-7.30 (m, 5H), 5.08 (s, 2H), 5.05-4.97 (m, 1H), 4.03-3.91 (m, 2H),0.84 (s, 9H), 0.03 (s, 6H);

LCMS (ESI): m/z 378.5 [M⁺+1]

Synthesis of (S)-2-((tert-butyldimethylsilyl) oxy)-1-(1, 3,4-oxadiazol-2-yl) ethan-1-amine (RR)

To a stirring solution of QQ (5 g, 13.2 mmol) in methanol (30 mL) wasadded 50% wet 10% Pd/C (1.2 g) and stirred under H₂ atmosphere (balloonpressure) for 2 h at RT. After completion of reaction, the reactionmixture was filtered through a pad of celite and triturated withmethanol (20 mL). The filtrate was concentrated under reduced pressureto afford RR (2 g, 63%) as thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.16 (s, 1H), 4.19-4.16 (m, 1H), 3.85-3.75(m, 2H), 2.11 (s, 2H), 0.77 (s, 9H), −0.04 (s, 6H)

LCMS (ESI): m/z 244.3 [M⁺+1]

Synthesis of 2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-5-isobutyryl-6-methyl-2, 5-diazaspiro [3.4] octan-1-one (C-38)

To a stirring solution of C-20 (500 mg, 1.81 mmol) in DCM (10 mL) wasadded DIPEA (0.94 mL, 5.43 mmol) at 0° C. After added isobutyrylchloride (578 mg, 5.43 mmol) at 0° C. and stirred at RT for 4 h.After consumption of the starting material (by TLC), the reactionmixture was diluted with water (10 mL). The separated organic layer wasdried over anhydrous Na₂SO₄ and concentrated under reduced pressure toafford crude compound which was purified by column chromatography byeluting 2% MeOH/DCM to obtained C-38 (150 mg, 24%) as pale yellow solid.

¹H-NMR: (400 MHz, CD₃OD): δ 8.82-8.77 (m, 2H), 7.39 (t, J=5.2 Hz, 1H),4.76 (d, J=8.0 Hz, 1H), 4.47-4.44 (m, 1H), 4.20-4.14 (m, 1H), 3.82-3.74(m, 1H), 3.51-3.40 (m, 1H), 2.78-2.72 (m, 1H), 2.46-2.37 (m, 1H),2.18-2.12 (m, 2H), 1.77-1.72 (m, 1H), 1.34-1.29 (m, 3H), 1.26-1.22 (m,3H), 1.06-1.02 (m, 6H)

LCMS (ESI): m/z 347.2 [M⁺+1];

HPLC: 99.7%

Synthesis of5-(3,3-dimethylbutanoyl)-2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-6-methyl-25-diazaspiro[3.4]octan-1-one(C-39)

To a stirring solution of C-20 (150 mg, 0.54 mmol) in DCM (10 mL) wasadded TEA (137 mg, 1.35 mmol) at 0° C. After added 3, 3-dimethylbutanoylchloride (87 mg, 0.65 mmol) at 0° C. and stirred at RT for 1 h. Afterconsumption of the starting material (by TLC), the reaction mixture wasdiluted with water (10 mL). The separated organic layer was dried overanhydrous Na₂SO₄ and concentrated under reduced pressure to afford crudecompound which was purified by column chromatography by eluting 2%MeOH/DCM to obtained C-39 (150 mg, 24%) as sticky solid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.80-8.75 (m, 2H), 7.43-7.37 (m, 1H),4.73-4.65 (m, 2H), 4.24-4.17 (m, 2H), 3.78-3.59 (m, 1H), 3.23-3.16 (m,1H), 2.36 (s, 2H), 2.32-2.08 (m, 3H), 1.58-1.50 (m, 1H), 1.20-1.08 (m,6H), 0.86 (s, 9H)

LCMS (ESI): m/z 375.49 [M⁺+1];

HPLC: 96.9%

Synthesis of cyclopentyl 2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-6-methyl-1-oxo-2, 5-diazaspiro [3.4] octane-5-carboxylate (C-40)

To a stirring solution of C-20 (150 mg, 0.54 mmol) in DCM (5 mL) wasadded TEA (0.23 mL, 1.63 mmol) at 0° C. After added cyclopentylchloroformate (88 mg, 0.59 mmol) at 0° C. and stirred at RT for 1 h.After consumption of the starting material (by TLC), the reactionmixture was diluted with water (10 mL). The separated organic layer waswashed with saturated brine solution (20 mL). The organic layer wasdried over anhydrous Na₂SO₄ and concentrated under reduced pressure toafford crude compound which was purified by column chromatography byeluting 3% MeOH/DCM to obtained C-40 (110 mg, 52%) as sticky solid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.81-8.76 (m, 2H), 7.45-7.42 (m, 1H),5.20-4.91 (m, 1H), 4.77-4.64 (m, 2H), 4.30-3.63 (m, 1H), 3.57-3.31 (m,3H), 2.16-1.86 (m, 4H), 1.77-1.32 (m, 8H), 1.18-1.08 (m, 6H)

LCMS (ESI): m/z 389.4 [M⁺+1];

UPLC: 97.6%

Synthesis of cyclohexyl 2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-6-methyl-1-oxo-2, 5-diazaspiro [3.4] octane-5-carboxylate (C-41)

To a stirring solution of C-20 (150 mg, 0.54 mmol) in DCM (5 mL) wasadded TEA (0.2 mL, 1.35 mmol) at 0° C. After added cyclohexylchloroformate (105 mg, 0.65 mmol) at 0° C. and stirred at RT for 1 h.After consumption of the starting material (by TLC), the reactionmixture was diluted with water (10 mL). The separated organic layer waswashed with citric acid solution (20 mL), saturated brine solution (20mL). The organic layer was dried over anhydrous Na₂SO₄ and concentratedunder reduced pressure to afford crude compound which was purified bycolumn chromatography by eluting 2% MeOH/DCM to obtained C-41 (90 mg,41%) as sticky solid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 8.82-8.78 (m, 2H), 7.48-7.41 (m, 1H),4.78-4.61 (m, 2H), 4.24-4.13 (m, 2H), 3.94-3.87 (m, 1H), 3.76-3.53 (m,2H), 2.27-1.99 (m, 3H), 1.97-1.92 (m, 1H), 1.56-1.32 (m, 10H), 1.29-1.10(m, 2H)

LCMS (ESI): m/z 403.5 [M⁺+1];

UPLC: 98.06%

Synthesis of 1-benzyl-N-((1S, 2R)-2-((tert-butyldimethylsilyl)oxy)-1-(1, 3, 4-oxadiazol-2-yl)propyl)-2-(hydroxymethyl)-5-methylpyrrolidine-2-carboxamide (C-42)

To a stirring solution of HH (2.5 g, 10.04 mmol) in DMF (15 mL) wereadded N, N-diisopropylethylamine (5.4 mL, 30.12 mmol), Int N (2.58 g,10.04 mmol) followed by HATU (4.57 g, 12.04 mmol) at 0° C. and stirredat RT for 16 h. After consumption of the starting material (by TLC), thereaction mixture was diluted with water (100 mL) and EtOAc (100 mL). Theorganic layer was washed with water (100 mL), saturated sodiumbicarbonate solution (50 mL) followed by brine solution (50 mL). Theorganic layer was dried over anhydrous Na₂SO₄, filtered and concentratedunder reduced pressure to afford crude compound which was purified bycolumn chromatography by eluting 20% EtOAc/n-hexane to obtained C-42(3.2 g, 65%) as brown syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.26 (s, 1H), 8.78 (d, J=10.0 Hz, 1H),7.30-7.21 (m, 5H), 5.26 (d, J=10.0 Hz, 1H), 4.43 (s, 2H), 4.05-4.02 (m,2H), 3.87-3.72 (m, 1H), 3.15-3.10 (m, 1H), 2.81-2.65 (m, 1H), 1.39-1.26(m, 4H), 1.25-1.16 (m, 6H), 0.05 (s, 6H)

LCMS (ESI): n/z 489.70 [M⁺+1]

Synthesis of 5-benzyl-2-((1S, 2R)-2-((tert-butyldimethylsilyl)oxy)-1-(1, 3, 4-oxadiazol-2-yl) propyl)-6-methyl-2, 5-diazaspiro[3.4]octan-1-one (C-43)

To a stirring solution of triphenylphosphine (4.02 g, 15.36 mmol) in dryTHF (30 mL) was added DIAD (2.48 g, 12.29 mmol) at RT and stirred for 15min. After added C-42 (3 g, 6.14 mmol) in THF (30 mL) was added dropwiseand the reaction mixture was stirred at RT for 4 h. After consumption ofthe starting material (by TLC), the reaction mixture was concentratedunder reduced pressure. The crude material was purified by columnchromatography by eluting 20% EtOAc/n-hexane to afford C-43 (1.3 g, 45%)as pale green syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.28 (s, 1H), 7.33-7.19 (m, 5H), 5.24-5.19(m, 1H), 4.73 (s, 2H), 4.47-4.41 (m, 1H), 4.02 (t, J=7.2 Hz, 1H), 3.68(s, 2H), 2.49-2.28 (m, 1H), 2.27-2.10 (m, 1H), 1.98-1.94 (m, 1H),1.47-1.40 (m, 1H), 1.23-1.15 (m, 6H), 0.87 (s, 9H), 0.02 (s, 6H)

LCMS (ESI): m/z 491.6 [M⁺+1]

Synthesis of 5-benzyl-2-((1S, 2R)-2-hydroxy-1-(1, 3, 4-oxadiazol-2-yl)propyl)-6-methyl-2, 5-diazaspiro [3.4] octan-1-one (C-44)

To a stirring solution of C-43 (1.3 g, 2.76 mmol) in dry THF (30 mL) wasadded TBAF (1M in THF) (4.13 mL, 4.14 mmol) at 0° C. and stirred for 2 hat RT. After consumption of the starting material (by TLC), the reactionmixture was concentrated under reduced pressure. The crude material wasdiluted with water (100 mL) and EtOAc (100 mL). The organic layer waswashed with water (100 mL), brine solution (50 mL). The organic layerwas dried over anhydrous Na₂SO₄, filtered and concentrated under reducedpressure to afford crude compound which was purified by columnchromatography by eluting 2% MeOH/CH₂Cl₂ to obtained C-44 (350 mg, 35%)as thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.18 (s, 1H), 7.34-7.16 (m, 5H), 5.21 (t,J=5.6 Hz, 1H), 5.01-4.83 (m, 1H), 4.22-4.18 (m, 1H), 3.88 (s, 2H),3.84-3.67 (m, 1H), 3.54-3.35 (m, 1H), 2.98-2.89 (m, 1H), 2.31-1.89 (m,3H), 1.45-1.41 (m, 1H), 1.39-1.14 (m, 3H), 0.90-0.81 (s, 3H)

LCMS (ESI): m/z 357.43 [M⁺+1]

HPLC: 92.46%

Synthesis of 5-benzyl-2-((1S, 2R)-2-hydroxy-1-(1, 3, 4-oxadiazol-2-yl)propyl)-6-methyl-2, 5-diazaspiro [3.4] octan-1-one (C-45)

To a stirring solution of C-44 (1.3 g, 2.76 mmol) in dry THF (30 mL) wasadded TBAF (1M in THF) (4.13 mL, 4.14 mmol) at 0° C. and stirred for 2 hat RT. After consumption of the starting material (by TLC), the reactionmixture was concentrated under reduced pressure. The crude material wasdiluted with water (100 mL) and EtOAc (100 mL). The organic layer waswashed with water (100 mL), brine solution (50 mL). The organic layerwas dried over anhydrous Na₂SO₄, filtered and concentrated under reducedpressure to afford crude compound which was purified by columnchromatography by eluting 2% MeOH/CH₂Cl₂ to obtained C-45 (350 mg, 35%)as thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.18 (s, 1H), 7.34-7.16 (m, 5H), 5.21 (t,J=5.6 Hz, 1H), 5.01-4.83 (m, 1H), 4.22-4.18 (m, 1H), 3.88 (s, 2H),3.84-3.67 (m, 1H), 3.54-3.35 (m, 1H), 2.98-2.89 (m, 1H), 2.31-1.89 (m,3H), 1.45-1.41 (m, 1H), 1.39-1.14 (m, 3H), 0.90-0.81 (s, 3H)

LCMS (ESI): m/z 357.43 [M⁺+1]

HPLC: 92.46%

Synthesis of2-((1S,2R)-2-hydroxy-1-(1,3,4-oxadiazol-2-yl)propyl)-3-methyl-2,5-diazaspiro[3.4]octan-1-one(C-46)

To a stirring solution of C-26 (800 mg, 2.18 mmol) in DCM (10 mL) wereadded BF₃OEt₂ (620 mg, 4.37 mmol), molecular sieves (150 mg) at 0° C.and stirred for 3 h at RT. After consumption of the starting material(by TLC), the reaction mixture was diluted with n-pentane (5 mL). Theprecipitated solid was filtered and filtrate was concentrated underreduced pressure. Obtained crude material was purified by silica gelcolumn chromatography eluting with 3% MeOH/DCM to afford C-46 (700 mg,crude) as yellow liquid.

Mass (ESI): m/z 267.3 [M⁺+1]

Synthesis of2-((1S,2R)-2-hydroxy-1-(1,3,4-oxadiazol-2-yl)propyl)-5-isobutyryl-3-methyl-2,5-diazaspiro[3.4]octan-1-one(C-47)

To a stirring solution of C-46 (700 mg (crude), 2.63 mmol) in DCM (10mL) was added TEA (0.53 mL, 3.94 mmol) at 0° C. After 10 min addedisobutyryl chloride (0.41 mL, 3.94 mmol) at 0° C. and stirred for 2 h atRT. After consumption of the starting material (by TLC), the reactionmixture was diluted with water (5 mL). The separated organic layer wasdried over anhydrous Na₂SO₄, filtered and concentrated under reducedpressure. Obtained crude material was purified by silica gel columnchromatography eluting with 3% MeOH/DCM followed by preparative HPLCpurification to afford C-47 (150 mg, 17%) as thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.27 (s, 1H), 5.20-5.17 (m, 1H), 4.87-4.85(m, 1H), 4.30-4.25 (m, 1H), 3.84-3.79 (m, 1H), 3.64-3.50 (m, 2H),2.72-2.66 (m, 1H), 2.13-2.09 (m, 1H), 2.01-1.79 (m, 3H), 1.37-1.19 (m,3H), 1.10-0.94 (m, 9H)

Mass (ESI): m/z 337.4 [M₊+1]

HPLC: 99.5%

Synthesis of 1-benzyl-N-((1S, 2R)-2-((tert-butyldimethylsilyl)oxy)-1-(1, 3, 4-oxadiazol-2-yl) propyl)-2-(1-hydroxyethyl)pyrrolidine-2-carboxamide (C-48)

To a stirring solution of LL (3.9 g, 15.66 mmol) in DMF (10 mL) wereadded N, N-diisopropylethylamine (8.16 mL, 46.98 mmol), Int-N (4.03 g,15.66 mmol) followed by HATU (7.14 g, 18.79 mmol) at 0° C. and stirredat RT for 16 h. After consumption of the starting material (by TLC), thereaction mixture was diluted with water (100 mL) and EtOAc (150 mL). Theseparated organic layer was washed with water (100 mL) followed by brinesolution (100 mL). The organic layer was dried over anhydrous Na₂SO₄ andconcentrated under reduced pressure to afford crude compound which waspurified by column chromatography by eluting 30% EtOAc/n-hexane toobtained C-48 (1.8 g, 24%) as yellow syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.25 (s, 1H), 8.74 (d, J=8.8 Hz, 1H),7.34-7.21 (m, 5H), 5.47-5.27 (m, 1H), 5.26-5.23 (m, 1H), 4.42-4.40 (m,1H), 4.13-4.01 (m, 2H), 3.82-3.77 (m, 1H), 2.70-2.60 (m, 2H), 1.99-1.82(m, 2H), 1.69-1.64 (m, 2H), 1.29-1.21 (m, 6H), 0.72 (s, 9H), −0.01 (s,3H), −0.02 (s, 3H);

LCMS (ESI): m/z 489.6 [M⁺+1]

Synthesis of 5-benzyl-2-((1S, 2R)-2-((tert-butyldimethylsilyl)oxy)-1-(1, 3, 4-oxadiazol-2-yl) propyl)-3-methyl-2, 5-diazaspiro[3.4]octan-1-one (C-49)

To a stirring solution of triphenylphosphine (2.41 g, 9.22 mmol) in dryTHF (20 mL) was added DIAD (1.49 g, 7.37 mmol) as portionwise andstirred for 15 min at RT. To this precipitated solution added C-48 (1.8g, 3.68 mmol) in dry THF (15 mL) dropwise at RT and stirred for 16 h.After consumption of the starting material (by TLC), the reactionmixture was concentrated under reduced pressure. The obtained solid wastriturated with 20% di ehylether/n-hexane (2×100 mL). The filterate wasconcentrated under reduced pressure to obtained crude compound which waspurified by silica gel column chromatography eluting 20% EtOAc/hexane toafford C-49 (400 mg, crude) as pale green syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.30 (s, 1H), 7.40-7.24 (m, 5H), 5.26-5.22(m, 1H), 4.80-4.73 (m, 2H), 4.62-4.48 (m, 1H), 4.15-4.00 (m, 1H),3.81-3.75 (m, 1H), 2.85 (t, J=8.0 Hz, 1H), 2.49-2.39 (m, 3H), 2.26-2.11(m, 1H), 1.71-1.64 (m, 3H), 1.25-1.15 (m, 3H), 0.78 (s, 9H), 0.02 (s,6H);

LCMS (ESI): m/z 471.6 [M⁺+1]

Synthesis of 5-benzyl-2-((1S, 2R)-2-hydroxy-1-(1, 3, 4-oxadiazol-2-yl)propyl)-3-methyl-2, 5-diazaspiro [3.4] octan-1-one (C-50)

To a stirring solution of C-49 (1.5 g, 3.19 mmol) in dry THF (30 mL) wasadded TBAF (1M in THF) (4.78 mL, 1.5 mmol) at 0° C. and stirred at RTfor 1 h. After consumption of the starting material (by TLC), thereaction mixture was concentrated under reduced pressure. The crudematerial was diluted with water (75 mL) and EtOAc (100 mL). The organiclayer was washed with water (2×50 mL), brine solution (2×50 mL). Theorganic layer was dried over anhydrous Na₂SO₄, filtered and concentratedunder reduced pressure to afford crude compound which was purified bycolumn chromatography by eluting 30% EtOAc/n-hexane to obtained C-50(500 mg, 44%) as white solid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.26 (s, 1H), 7.37-7.22 (m, 5H), 5.29-5.21(m, 1H), 4.96-4.75 (m, 1H), 4.33-4.02 (m, 1H), 4.00-3.94 (m, 2H),3.77-3.70 (m, 2H), 2.91-2.80 (m, 1H), 2.42-2.38 (m, 1H), 2.16-2.08 (m,2H), 1.80-1.63 (m, 1H), 1.30 (d, J=6.4 Hz, 3H), 1.12 (d, J=6.0 Hz, 3H);

LCMS (ESI): m/z 357.43 [M⁺+1]

HPLC: 99.61%

Synthesis of2-((1S,2R)-2-hydroxy-1-(1,3,4-oxadiazol-2-yl)propyl)-3-methyl-2,5-diazaspiro[3.4]octan-1-one(C-51)

To a stirring solution of compound C-50 (420 mg, 1.17 mmol) in methanol(10 mL) was added 10% Pd/C (400 mg) under N₂ atmosphere. The reactionmixture was stirred under H₂ atmosphere at RT for 3 h. After consumptionof the starting material (by TLC), the reaction mixture was filteredthrough a pad of celite and the pad was washed with methanol (10 mL).Obtained filtrate was concentrated under reduced pressure to afford C-51(220 mg, 70%) as an off-white solid.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.26 (s, 1H), 5.25-5.17 (m, 1H), 4.89-4.69(m, 1H), 4.28-4.21 (m, 1H), 3.84-3.56 (m, 2H), 2.95-2.79 (m, 2H),1.99-1.84 (m, 2H), 1.77-1.59 (m, 2H), 1.24-1.10 (m, 6H):

LCMS (ESI): m/z 260 [M⁺+1]

HPLC: 97.94%

Synthesis of 2-(2-hydroxy-1-(pyrimidin-2-yl) propyl)-3, 6-dimethyl-2,5-diazaspiro [3.4]octan-1-one (C-52)

To a stirring solution of C-33 (500 mg, 1.72 mmol) in dry DCM (10 mL)was added DIPEA (556 mg, 4.31 mmol) followed by isobutyryl chloride (273mg, 2.58 mmol) slowly at 0° C. and stirred at RT for 3 h. Afterconsumption of the starting material (by TLC), the reaction mixture wasconcentrated under reduced pressure. The crude residue was diluted withwater (10 mL). The separated organic layer was washed with brinesolution (20 mL). The separated organic layer was dried over anhydrousNa₂SO₄ and concentrated under reduced pressure to afford crude compoundwhich was purified by column chromatography by eluting 1% MeOH/DCM toobtained C-52 (90 mg, 14.5%) as pale brown syrup.

¹H-NMR: (400 MHz, CD₃OD): δ 8.81-8.77 (m, 2H), 7.41-7.37 (m, 1H),4.63-4.53 (m, 2H), 4.24-4.18 (m, 2H), 2.87-2.72 (m, 1H), 2.33-2.27 (m,1H), 2.16-2.08 (m, 2H), 1.73-1.69 (m, 1H), 1.40-1.35 (m, 3H), 1.28-1.20(m, 3H), 1.15-1.02 (m, 9H);

LCMS (ESI): m/z 361.3 [M⁺+1];

HPLC: 94.85%

Synthesis of ethyl 2-(1-hydroxyethyl)-5-methylpyrrolidine-2-carboxylate(C-53)

To a stirring solution of CC (5 g, 16.61 mmol) in CH₂Cl₂ (50 mL) wasadded TFA (6.34 mL, 83.05 mmol) at 0° C. After being stirred at RT for 4h, the reaction mixture was concentrated under reduced pressure toafford crude compound which was triturated with n-pentane (50 mL) toobtained C-53 (5 g, crude) as black syrup was directly taken for thenext step without further purification.

¹H-NMR: (500 MHz, DMSO-d₆): δ 8.71 (br s, 1H), 4.29-4.25 (m, 1H),4.25-4.21 (m, 3H), 4.14-4.03 (m, 1H), 2.31-2.10 (m, 4H), 1.36-1.18 (m,6H), 1.18-1.10 (m, 3H)

LCMS (ESI): m/z 202.38 [M⁺+1]

Synthesis of ethyl1-benzyl-2-(1-hydroxyethyl)-5-methylpyrrolidine-2-carboxylate (C-54:C-54-A & C-54-B)

To a stirring solution of C-53 (5 g (crude), 15.87 mmol) in acetonitrile(0.50 mL) was added K₂CO₃ (6.57 g, 47.61 mmol) followed by benzylbromide (2.82 mL, 23.80 mmol) at RT and stirred for 16 h. Aftercompletion of the reaction, diluted the reaction mixture with EtOAc (150mL) and water (75 mL). The organic layer was washed with brine solution(2×150 mL). The organic layer was dried over anhydrous Na₂SO₄ andconcentrated under vacuum. Obtained crude material was purified bycolumn chromatography eluting with 10% EtOAc/Hexane to afford 1 g ofC-54-A and 1.2 g of C-54-B (separated isomers of C-54) (48%) as yellowsyrups.

¹H-NMR (C-54-A isomer): (500 MHz, DMSO-d₆): δ 7.35-7.14 (m, 5H), 4.55(s, 2H), 4.24-4.19 (m, 1H), 4.13-4.02 (m, 2H), 3.96-3.91 (m, 2H),2.14-1.99 (m, 1H), 1.95-1.82 (m, 3H), 1.25-1.18 (m, 3H), 1.10-1.06 (m,3H), 0.75 (d, J=6.0 Hz, 3H)

LCMS (ESI): m/z 292.3 [M⁺+1]

¹H-NMR (C-54-B isomer): (500 MHz, DMSO-d₆): δ 7.34-7.18 (m, 5H), 5.15(t, J=5.5 Hz, 2H), 4.63-4.49 (m, 1H), 4.24-4.22 (m, 1H), 4.10-4.06 (m,2H), 3.18-3.10 (m, 1H), 2.39-2.33 (m, 1H), 1.89-1.83 (m, 3H), 1.42-1.39(m, 3H), 1.20-1.13 (m, 3H), 0.76 (d, J=6.0 Hz, 3H)

LCMS (ESI): m/z 292.3 [M⁺+1]

Synthesis of1-benzyl-2-(1-hydroxyethyl)-5-methylpyrrolidine-2-carboxylic Acid(C-55-A)

To a stirring solution of C-54-A (1 g, 3.43 mmol) in MeOH/THF/H₂O (5mL/5 mL/5 mL) were added NaOH (206 mg, 5.14 mmol) at 0° C. The reactionmixture was heated to 90° C. for 6 h. After consumption of the startingmaterial (by TLC), the solvent was evaporated under reduced pressure.The aqueous layer was washed with di ethylether (50 mL). The separatedaqueous layer was acidified by using 2N HCl (pH˜3). The aqueous layerwas extracted with 10% MeOH/DCM (2×100 mL). The combined organic layerwas dried over Na₂SO₄ and concentrated under reduced pressure to affordcompound C-55-A (1 g, crude) as yellow solid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.84 (s, 1H), 7.60 (d, J=6.8 Hz, 2H),7.50-7.30 (m, 3H), 5.12 (br s, 1H), 4.48-4.35 (m, 2H), 4.09-3.81 (m,1H), 2.39-2.22 (m, 2H), 2.16-2.08 (m, 2H), 1.66-1.57 (m, 1H), 1.16-0.86(m, 6H);

LCMS (ESI): m/z 264.3 [M⁺+1]

Synthesis of1-benzyl-2-(1-hydroxyethyl)-5-methylpyrrolidine-2-carboxylic Acid(C-55-B)

To a stirring solution of C-54-B (1.2 g, 4.12 mmol) in MeOH/THF/H₂O (5mL/5 mL/5 mL) were added NaOH (206 mg, 5.14 mmol) at 0° C. The reactionmixture was heated to 90° C. for 6 h. After consumption of the startingmaterial (by TLC), the solvent was evaporated under reduced pressure.The aqueous layer was washed with di ethylether (50 mL). The separatedaqueous layer was acidified by using 2N HCl (pH˜3). The aqueous layerwas extracted with 10% MeOH/DCM (2×100 mL). The combined organic layerwas dried over Na₂SO₄ and concentrated under reduced pressure to affordcompound C-55-B (780 mg) as yellow solid.

¹H-NMR (C-55-B isomer): (400 MHz, DMSO-d₆): δ 9.14 (s, 1H), 7.64 (d,J=6.0 Hz, 2H), 7.49-7.44 (m, 3H), 4.99 (d, J=12.8 Hz, 1H), 4.48 (s, 2H),4.45-4.41 (m, 1H), 3.98-3.93 (m, 1H), 2.32-2.13 (m, 4H), 1.40 (d, J=6.4Hz, 3H), 0.76 (d, J=6.4 Hz, 3H);

LCMS (ESI): m/z 272.4 [M⁺−1]

Synthesis of 1-benzyl-N-((1S, 2R)-2-((tert-butyldimethylsilyl)oxy)-1-(1,3, 4-oxadiazol-2-yl)propyl)-2-(1-hydroxyethyl)-5-methylpyrrolidine-2-carboxamide (C-56)

To a stirring solution of C-55 (1.78 g, 6.75 mmol)(mixture of twoisomers C-55-A & C-55-B) in DMF (10 mL) were added N,N-diisopropylethylamine (3.51 mL, 20.27 mmol), Int N (1.74 g, 6.75 mmol)followed by HATU (3.08 g, 8.11 mmol) at 0° C. and stirred at RT for 16h. After consumption of the starting material (by TLC), the reactionmixture was diluted with water (100 mL) and EtOAc (150 mL). Theseparated organic layer was washed with water (100 mL) followed by brinesolution (100 mL). The organic layer was dried over anhydrous Na₂SO₄ andconcentrated under reduced pressure to afford crude compound which waspurified by column chromatography by eluting 30% EtOAc/n-hexane toobtained C-56 (1.2 g, 35%) as brown syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.31 (s, 1H), 8.42 (s, 1H), 7.44-7.19 (m,5H), 5.26-4.95 (m, 1H), 4.36 (s, 2H), 4.33-4.18 (m, 2H), 4.09-3.94 (m,1H), 2.32-2.22 (m, 1H), 2.15-1.86 (m, 4H), 1.37-1.05 (m, 9H), 0.85 (s,9H), 0.04 (s, 6H);

LCMS (ESI): m/z 503.7 [M⁺+1]

Synthesis of 1-benzyl-N-((1S, 2R)-2-((tert-butyldimethylsilyl)oxy)-1-(1, 3, 4-oxadiazol-2-yl)propyl)-2-(1-hydroxyethyl)-5-methylpyrrolidine-2-carboxamide (C-57)

To a stirring solution of triphenylphosphine (1.56 g, 5.97 mmol) in dryTHF (15 mL) was added DIAD (967 mg, 4.78 mmol) as portionwise andstirred for 15 min at RT. To this precipitated solution added C-56 (1.2g, 2.39 mmol) in dry THF (15 mL) slowly at RT and stirred for 4 h. Afterconsumption of the starting material (by TLC), the reaction mixture wasconcentrated under reduced pressure. The obtained solid was trituratedwith 20% di ehylether/n-hexane (2×100 mL). The filterate wasconcentrated under reduced pressure to obtained crude compound which waspurified by silica gel column chromatography eluting 20% EtOAc/hexane toafford C-57 (400 mg, crude) as brown thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.27 (s, 1H), 7.43-7.18 (m, 5H), 4.91-4.73(m, 2H), 4.42-4.02 (m, 2H), 3.96-3.75 (m, 2H), 2.11-1.84 (m, 3H),1.42-1.37 (m, 1H), 1.35 (d, J=6.0 Hz, 3H), 1.23-1.11 (m, 6H), 0.70 (s,9H), 0.02 (s, 6H);

LCMS (ESI): m/z 485.7 [M⁺+1]

Synthesis of 5-benzyl-2-((1S, 2R)-2-hydroxy-1-(1, 3, 4-oxadiazol-2-yl)propyl)-3, 6-dimethyl-2, 5-diazaspiro [3.4]octan-1-one (C-58)

To a stirring solution of C-57 (400 mg, 0.82 mmol) in dry THF (5 mL) wasadded TBAF (1M in THF) (1.23 mL, 1.5 mmol) at 0° C. and stirred for 1 hat RT. After consumption of the starting material (by TLC), the reactionmixture was concentrated under reduced pressure. The crude material wasdiluted with water (30 mL) and EtOAc (50 mL). The organic layer waswashed with water (2×20 mL), brine solution (2×20 mL). The organic layerwas dried over anhydrous Na₂SO₄, filtered and concentrated under reducedpressure to afford crude compound which was purified by columnchromatography by eluting 30% EtOAc/n-hexane to obtained C-58 (90 mg,29%) as thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.25 (s, 1H), 7.41-7.18 (m, 5H), 5.28-5.17(m, 1H), 4.97-4.62 (m, 1H), 4.43-4.20 (m, 1H), 4.02-3.89 (m, 1H),3.77-3.64 (m, 2H), 3.20-3.01 (m, 1H), 2.09-1.80 (m, 3H), 1.44-1.40 (m,1H), 1.38-1.30 (m, 1H), 1.28-1.21 (m, 4H), 1.19-1.06 (m, 1H), 0.90-0.85(m, 3H)

LCMS (ESI): m/z 371.45 [M⁺+1]

HPLC: 94.53%

Synthesis of tert-butyl2-(((S)-2-(tert-butyldimethylsilyl)oxy)-1-(1,3,4-oxadiazol-2-yl)ethyl)carbamoyl)-2-(hydroxymethyl)-5-methylpyrrolidine-1-carboxylate(C-59)

To a stirring solution of Z (2 g, 7.72 mmol) in DCM (25 mL) were addedN, N-diisopropylethylamine (4 mL, 23.16 mmol), Int RR (2.06 g, 8.49mmol), followed by HATU (3.52 g, 9.26 mmol) at 0° C. and stirred at RTfor 16 h. After consumption of the starting material (by TLC), thereaction mixture was diluted with water (50 mL). The separated organiclayer was washed with citric acid solution (1×50 mL), saturated brinesolution (1×50 mL). The separated organic layer was dried over anhydrousNa₂SO₄ and concentrated under reduced pressure to afford crude compoundwhich was purified by column chromatography by eluting 50%EtOAc/n-hexane to obtained C-59 (2.5 g, 67%) as thick syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.25 (s, 1H), 7.99 (d, J=3.5 Hz, 1H),5.29-5.26 (m, 2H), 4.05-3.99 (m, 3H), 3.57-3.54 (m, 2H), 2.28-1.91 (m,3H), 1.42 (s, 9H), 1.38-1.31 (m, 1H), 1.18-1.11 (m, 3H), 0.84 (s, 9H),−0.01 (s, 6H)

Synthesis of tert-butyl2-((S)-2-((tert-butyldimethylsilyl)oxy)-1-(1,3,4-oxadiazol-2-yl)ethyl)-6-methyl-1-oxo-2,5-diazaspiro[3.4]octane-5-carboxylate(C-60)

To a stirring solution of triphenylphosphine (2.7 g, 10.33 mmol) in dryTHF (20 mL) was added DTAD (2.37 g, 10.33 mmol) at RT and stirred for 15min. After added C-59 (2.5 g, 5.16 mmol) and the reaction mixture wasstirred at RT for 3 h. After consumption of the starting material (byTLC), the reaction mixture was concentrated under reduced pressure. Thecrude material was triturated with di ethylether/n-pentane and obtainedsolid was filtered. The filtrate was evaporated, purified by silica gelcolumn chromatography eluting 30% EtOAc/hexane to afford C-60 (2.1 g,87.5%) as yellow liquid.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.32 (s, 1H), 5.19-5.08 (m, 1H), 4.12-3.87(m, 3H), 3.58-3.45 (m, 2H), 1.56-1.42 (m, 3H), 1.40 (s, 9H), 1.38-1.20(m, 1H), 1.18-1.12 (m, 3H), 0.80 (s, 9H), 0.02 (s, 6H)

Synthesis of tert-butyl 2-(2-hydroxy-1-(pyrimidin-2-yl)propyl)-6-methyl-1-oxo-2, 5-diazaspiro [3.4] octane-5-carboxylate (C-61)

To a stirring solution of C-60 (2.1 g, 4.5 mmol) in dry THF (10 mL) wasadded TBAF (2.34 g, 9 mmol) at 0° C. The reaction mixture was stirred atRT for 3 h. After consumption of the starting material (by TLC), thereaction mixture was concentrated under reduced pressure. The crudematerial was purified by silica gel column chromatography eluting 2%MeOH/DCM to afford C-61 (800 mg, 51%) as thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.25 (s, 1H), 5.33-5.08 (m, 2H), 3.94-3.75(m, 3H), 3.67-3.51 (m, 1H), 3.39-3.31 (m, 1H), 2.13-1.93 (m, 3H),1.55-1.50 (m, 1H), 1.40 (s, 9H), 1.15-1.08 (m, 3H),

LCMS (ESI): m/z 353.3 [M⁺+1];

HPLC: 96.6%

Synthesis of tert-butyl 2-(((S)-2-((tert-butyldimethylsilyl) oxy)-1-(1,3, 4-oxadiazol-2-yl) ethyl) carbamoyl)-2-(1-hydroxyethyl)pyrrolidine-1-carboxylate (C-62)

1 To a stirring solution of BB (3.5 g, 13.51 mmol) in CH₂Cl₂ (40 mL)were added DIPEA (7.0 mL, 40.53 mmol), RR (3.2 g, 13.51 mmol), HATU(0.5.6 g, 14.85 mmol) at 0° C. and stirred for 12 h. After consumptionof the starting material (by TLC), the reaction mixture was diluted withwater (30 mL) and extracted with CH₂Cl₂ (2×100 mL). The separatedorganic layer was washed with brine, dried over anhydrous Na₂SO₄,filtered and concentrated under reduced pressure to afford C-62 (3.5 g.crude) as colorless liquid.

Mass (ESI): m/z 485.67 [M⁺+1]

Synthesis of tert-butyl 2-((S)-2-((tert-butyldimethylsilyl) oxy)-1-(1,3, 4-oxadiazol-2-yl) ethyl)-1-methyl-3-oxo-2, 5-diazaspiro [3.4]octane-5-carboxylate (C-63)

To a stirring solution of triphenylphosphine (2.03 g, 7.75 mmol) in THF(25 mL) was added DIAD (1.59 g, 7.75 mmol) at 0° C. and stirred for 30min. C-62 (1.5 g, 3.10 mmol) in THF (10 mL) was added dropwise and thereaction mixture was stirred at RT for 16 h. After consumption of thestarting material (by TLC), the reaction was concentrated under reducedpressure. The crude material was purified by silica gel columnchromatography eluting 30% EtOAc/hexane to afford C-63 (600 mg, 43%) aspale yellow liquid.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.24 (s, 1H), 5.93-4.75 (m, 2H), 4.16-3.91(m, 2H), 3.78-3.70 (m, 2H), 3.91 (d, J=7.0 Hz, 1H), 3.77 (d, J=7.0 Hz,2H), 3.44-3.34 (m, 4H), 2.01-1.91 (m, 2H), 1.85-1.68 (m, 6H), 2.11-1.68(m, 4H), 1.40 (s, 9H), 1.38-1.18 (m, 3H), 0.80 (s, 9H), −0.02 (s, 6H).

Mass (ESI): m/z 467.6 [M⁺+1]

Synthesis of tert-butyl 2-((S)-2-hydroxy-1-(1, 3, 4-oxadiazol-2-yl)ethyl)-1-methyl-3-oxo-2, 5-diazaspiro [3.4] octane-5-carboxylate (C-64)

To a stirring solution of C-63 (0.5 g, 1.07 mmol) in dry THF (10 mL) wasadded TBAF (1M in THF) (1.07 mL, 1.91 mmol) at 0° C. and stirred at RTfor 3 h. After consumption of the starting material (by TLC), thereaction mixture was concentrated under reduced pressure. The crudematerial was diluted with water (15 mL) and EtOAc (30 mL). The organiclayer was washed with water (2×15 mL), brine solution (2×10 mL). Theorganic layer was dried over anhydrous Na₂SO₄, filtered and concentratedunder reduced pressure to afford crude compound which was purified bycolumn chromatography by eluting 70% EtOAc/n-hexane to obtained C-64 (95mg, 25%) as white solid.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.26 (s, 1H), 5.32-5.04 (m, 2H), 3.98-3.72(m, 4H), 2.14-1.66 (m, 4H), 1.39 (s, 1H), 1.37-1.09 (m, 3H)

LCMS (ESI): m/z 353.3 [M⁺+1]

HPLC: 88.9%

Synthesis of tert-butyl2-(((S)-2-((tert-butyldimethylsilyl)oxy)-1-(1,3,4-oxadiazol-2-yl)ethyl)carbamoyl)-2-(1-hydroxyethyl)-5-methylpyrrolidine-1-carboxylate(C-65)

To a stirring solution of DD (1 g, 3.66 mmol) in DCM (20 mL) were addedN, N-diisopropylethylamine (1.9 mL, 10.98 mmol), Int RR (978 mg, 4.02mmol), EDCI (1.04 g, 5.49 mmol) followed by HOBT (741 mg, 5.49 mmol) at0° C. and stirred at RT for 16 h. After consumption of the startingmaterial (by TLC), the reaction mixture was diluted with water (20 mL).The separated organic layer was washed with brine solution (30 mL). Theseparated organic layer was dried over anhydrous Na₂SO₄ and concentratedunder reduced pressure to afford crude compound which was purified bycolumn chromatography by eluting 40% EtOAc/n-hexane to obtained C-65(700 mg, 38.4%) as thick syrup.

¹H-NMR: (500 MHz, DMSO-d₆): δ 9.26 (s, 1H), 5.22-5.18 (m, 1H), 4.58-4.50(m, 2H), 4.04-3.92 (m, 3H), 1.47-1.41 (m, 1H), 1.38 (s, 9H), 1.29-1.16(m, 6H), 1.14-1.11 (m, 3H), 0.83 (s, 9H), −0.02 (s, 6H);

LCMS (ESI): m/z 523.6 [M⁺+1]

Synthesis of tert-butyl2-((S)-2-((tert-butyldimethylsilyl)oxy)-1-(1,3,4-oxadiazol-2-yl)ethyl)-1,6-dimethyl-3-oxo-2,5-diazaspiro[3.4]octane-5-carboxylate(C-66)

To a stirring solution of triphenylphosphine (736 mg, 2.81 mmol) in dryTHF (10 mL) was added DTAD (646 mg, 2.81 mmol) as portionwise andstirred for 15 min at RT. To this precipitated solution added C-65 (700mg, 1.4 mmol) in dry THF (15 mL) slowly at RT and stirred for 6 h. Afterconsumption of the starting material (by TLC), the reaction mixture wasconcentrated under reduced pressure. The crude material was trituratedwith n-pentane (20 mL)/di ehylether (20 mL). The filterate wasconcentrated under reduced pressure to obtained crude compound which waspurified by silica gel column chromatography eluting 25% EtOAc/hexane toafford C-66 (430 mg, 64%) as thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.25 (s, 1H), 5.20-5.16 (m, 1H), 4.16-4.00(m, 1H), 3.88-3.70 (m, 3H), 2.17-2.08 (m, 1H), 1.96-1.85 (m, 1H), 1.40(s, 9H), 1.26-1.07 (m, 8H), 0.82 (s, 9H), −0.02 (s, 6H);

LCMS (ESI): m/z 505.5 [M⁺+1]

Synthesis of tert-butyl 2-((S)-2-hydroxy-1-(1, 3, 4-oxadiazol-2-yl)ethyl)-1, 6-dimethyl-3-oxo-2, 5-diazaspiro [3.4] octane-5-carboxylate(C-67)

To a stirring solution of C-66 (430 mg, 0.89 mmol) in dry THF (10 mL)was added TBAF (1.8 mL, 1.79 mmol) slowly at 0° C. and stirred at RT for6 h. After consumption of the starting material (by TLC), the reactionmixture was concentrated under reduced pressure. The crude residue waspurified with column chromatography by eluting 90% EtOAc/n-hexane toafford C-67 (80 mg, 24.5%) as thick syrup.

¹H-NMR: (400 MHz, DMSO-d₆): δ 9.25 (s, 1H), 5.20-5.03 (m, 2H), 3.98-3.72(m, 3H), 2.50-1.85 (m, 3H), 1.56-1.50 (m, 1H), 1.40 (s, 9H), 1.27-1.11(m, 6H)

LCMS (ESI): m/z 291.3 [M⁺+1];

HPLC: 91.4%

Example 3—[³H] MK-801 Binding Assay

Methods

Assays were conducted as described in Moskal et al. (Moskal, J. R., Kuo.A. G., Weiss, C., Wood. P. L., O'Connor Hanson, A., Kelso, S., Harris,R. B., Disterhoft, J. F., 2005. GLYX-13: a monoclonal antibody-derivedpeptide that acts as an N-methyl-D-aspartate receptor modulator.Neuropharmacology, 49, 1077-87) The potentiation of [³H]MK-801 binding(5 nM; 22.5 Ci/mmol) to well washed rat cortical membranes (200 μg) wasmeasured under non-equilibrium conditions (15 min @ 25° C.) in thepresence of increasing concentrations of test compounds and 50 μMglutamate. Zero levels were determined in the absence of any glycineligand and in the presence of 30 μM 5,7 DCKA. Maximal stimulation wasmeasured in the presence of 1 mM glycine, and 50 μM glutamate waspresent in all samples. The facilitation of [³H]MK-801 binding by testscompounds was calculated by using a 3 parameter log agonist vs. responseequation (Graph pad Prism. USA) and potency (EC₅₀, expressed in pM) andmaximal activity (% maximal stimulation) were calculated for the testcompound.

Results

The potency and maximal activity for Compound Y is 288 pM, 14%.

TABLE 2 Additional Biological Data Unified Unified Activity UnifiedMK-801 Activity Data: Unified Activity Glycine Unified Unified Data:LTP, Porsolt Activity Unified Data: Site Activity Data: Activity Data:Significant Floating Data: Activity Porsolt Binding LTP LTP (S) or Non-Time Porsolt Data: Time Post Assay: Rat Augmentation Concentrationsignificant Inhibition Dose Porsolt Dose Cortex Compound (Percent) (uM)(NS) (Percent) (mg/kg) Dose, route (Hours) EC50 (M) C-4 140 1 S 92 3 IV1 2.879E−10 C-15 120 1 S 86 1 PO 1

TABLE 3 Additional Biological Data MK-801 Glycine Site Binding Assay:Rat LTP: LTP LTP: LTP LTP: LTP Cortex Augmentation ConcentrationSignificance, Compound EC50 (M) (%) (uM) S or NS C-33 7.129E−12 30 1 NSC-19 1.096E−10 110 1 S C-26  7.23E−13 65 1 NS C-29 1.353E−10 125 1 S

EQUIVALENTS

Those skilled in the art will recognize, or be able to ascertain usingno more than routine experimentation, many equivalents to the specificembodiments of the invention described herein. Such equivalents areintended to be encompassed by the following claims.

INCORPORATION BY REFERENCE

The entire contents of all patents, published patent applications,websites, and other references cited herein are hereby expresslyincorporated herein in their entireties by reference.

1.-4. (canceled)
 5. A compound represented by formula I:

or a stereoisomer and/or a pharmaceutically acceptable salt thereof,wherein: R_(b) is H; R₁ is H; R₂ is H; R₃ is —C(O)R₃₂, wherein R₃₂ is—CH₃; R₅ has the formula —CH(CHR₄R₆)—X, wherein X is 1,3,4-oxadiazolyl;R₄ is methyl; and R₆ is hydroxyl.
 6. A pharmaceutical composition, thepharmaceutical composition comprising: a compound represented by formulaI:

or a stereoisomer and/or a pharmaceutically acceptable salt thereof,wherein: R_(b) is H; R₁ is H; R₂ is H; R₃ is —C(O)R₃₂, wherein R₃₂ is—CH₃; R₅ has the formula —CH(CHR₄R₆)—X, wherein X is 1,3,4-oxadiazolyl;R₄ is methyl; and R₆ is hydroxyl; and a pharmaceutically acceptableexcipient.